Search

Introduction: Spermatogenesis is the primary process of sperm production occuring in seminiferous tubules. Spermatogonial stem cells (SSCs) have the ability for self-renewal, differentiation, and the transmission of genetic information to subsequent generations. KLF4 and POU5F1 are transcription factors expressed in a wide range of tissues and play key roles in such processes as apoptosis, differentiation, proliferation, and cellular development. The present study aimed to assess the expression levels of KLF4 and POU5F1 genes in mouse embryonic stem cells (mESCs), spermatogonial stem cells (SSCs), embryonic-like stem cells (ES-like), and testicular cells, and identify the signaling pathways associated with them in the process of spermatogenesis. Materials & Methods: In this experimental study, spermatogonial cells were extracted from mouse testes using enzymatic digestion method and cultured in GSC medium containing FGF, EGF, and GDNF. Thereafter, the expression of KLF4 and POU5F1 genes was examined in mESCs, SSCs, ES-like, and testicular cells was investigated using immunocytochemistry, immunohistochemistry, and reverse transcription-polymerase chain reaction methods, and protein-protein interactions and signaling pathways were evaluated using bioinformatics methods. Results: The KLF4 and POU5F1 genes exhibited positive expression in ES-like cells and testicular cells. The assessment of KLF4 mRNA and POU5F1 mRNA expression levels demonstrated that KLF4 expression is higher in mESCs and ES-like cells compared to other cells, while POU5F1 expression is higher in SSCs. Both KLF4 and POU5F1 are considered essential and powerful genes that share a common class and function. Conclusion: The findings of this study indicated that KLF4 and POU5F1 play crucial roles in the proper development of sperm and are present in various types of cells, including mESCs, SSCs, ES-like cells, and testicular cells. These factors are key components of sexual stem cells and contribute to stem cell proliferation, making them potential diagnostic markers for these cell lines. [ABSTRACT FROM AUTHOR]

Background and purpose: Chronic Lymphocytic Leukemia (CLL) is a lymphoproliferative disorder characterized by the clonal and malignant proliferation of mature B-cells within the peripheral blood, bone marrow, and secondary lymphoid tissues. In chronic infections and cancers, T-cells become exhausted due to continuous antigen exposure. This uncontrolled tumor cell growth creates an environment that lacks sufficient oxygen, nutrients, and physical space for normal cells, referred to as the tumor microenvironment. The tumor microenvironment employs multiple mechanisms to suppress the immune system, affecting metabolic pathways in both T-cells and tumor cells. Consequently, inhibitory receptors on T-cells and their ligands on tumor cells are upregulated, leading to reduced proliferation, cytotoxicity, and production of effector cytokines, such as interferon-gamma, by T-cells. T-cell exhaustion presents a therapeutic challenge in treating many malignancies, including CLL. In CLL, T-cell exhaustion is observed through the simultaneous upregulation of inhibitory receptors. Given that current therapies for CLL are often associated with side effects, resistance, and relapse, studying transcript changes may assist in designing more effective treatment strategies. This study investigates the transcriptome profile of CD8+ T-cells in CLL patients compared to healthy individuals using bioinformatics platforms. Materials and methods: The keywords used in this study were CLL, CD8, and human. Raw data were initially searched in the SRA database using these keywords. Left and right reads were merged using CLC Genomics Workbench version 21. Following quality control of the data, sequences were assembled using human genome reference data, mRNA sequences, and coding sequences (CDS). Gene expression related to CD8+ T-cell exhaustion was then compared between CLL patients and healthy individuals. The analysis results were presented in heatmap and volcano plot formats. Results: In this study, genes involved in CD8+ T-cell exhaustion and differential expression were identified by comparing 11 CLL patients with 11 healthy individuals. The expression levels of 59 selected genes were displayed in volcano plot and heatmap formats. Expression changes in 42 genes were statistically significant (FDR P

Introduction: Breast cancer, which is one of the most common cancers with high mortality in women, has always been the focus of researchers, and every day, scientists are trying to identify mechanisms, genes, and medicines related to this disease. Nowadays, bioinformatics methods are used to identify and repurpose drugs for the treatment of diseases, especially cancer. Material & Methods: In this study, bioinformatics and biological network analysis were used to identify candidate drugs for breast cancer treatment. In this regard, analysis of the protein interaction network and drug-gene network were employed. The needed data were collected from the GEO database with the access code GSE54002. For the selected data set, genes with significant expression changes between two groups of healthy people and people with breast cancer cases were selected and considered primary genes. Thereafter, the protein-protein interaction network was constructed using the STRING database, and a significant gene module was obtained from the network. Following that, gene ontology studies and biological pathways were conducted. Next, the drug-gene network was constructed to identify drugs that target module genes and were introduced as essential drugs for the treatment of breast cancer. Cytoscape software and STRING and OncoDB databases were used to reconstruct and analyze the networks. Results: After analyzing the protein-protein interaction network and the drug-gene network, three important drugs that target the genes of the modules were identified and introduced as candidate drugs for the treatment of breast cancer. These drugs were RG-1530, R-406, and GW441756x. Discussion & Conclusion: The obtained results demonstrated that the introduced drugs (RG-1530, R-406, and GW441756x) can be effective in the treatment of breast cancer

Introduction: Breast cancer, which is one of the most common cancers with high mortality in women, has always been the focus of researchers, and every day, scientists are trying to identify mechanisms, genes, and medicines related to this disease. Nowadays, bioinformatics methods are used to identify and repurpose drugs for the treatment of diseases, especially cancer. Material & Methods: In this study, bioinformatics and biological network analysis were used to identify candidate drugs for breast cancer treatment. In this regard, analysis of the protein interaction network and drug-gene network were employed. The needed data were collected from the GEO database with the access code GSE54002. For the selected data set, genes with significant expression changes between two groups of healthy people and people with breast cancer cases were selected and considered primary genes. Thereafter, the protein-protein interaction network was constructed using the STRING database, and a significant gene module was obtained from the network. Following that, gene ontology studies and biological pathways were conducted. Next, the drug-gene network was constructed to identify drugs that target module genes and were introduced as essential drugs for the treatment of breast cancer. Cytoscape software and STRING and OncoDB databases were used to reconstruct and analyze the networks. Results: After analyzing the protein-protein interaction network and the drug-gene network, three important drugs that target the genes of the modules were identified and introduced as candidate drugs for the treatment of breast cancer. These drugs were RG1530, R-406, and GW441756x. Discussion & Conclusion: The obtained results demonstrated that the introduced drugs (RG-1530, R-406, and GW441756x) can be effective in the treatment of breast cancer. [ABSTRACT FROM AUTHOR]

Introduction: Brucellosis is one of the most common zoonotic diseases worldwide, posing a significant threat to both human health and the livestock industry, causing substantial economic losses. Today, multi-epitope antigens are widely used in serological diagnostic methods, especially ELISA, due to their high sensitivity and specificity in diagnostic efficiency and lower production costs compared to single antigens. Therefore, this study aimed to design a bioinformatic multi-epitope antigen for the diagnosis of human brucellosis. Methods: To design the multi-epitope antigen, Omp16, Omp25, and BP26 antigens were utilized. After identifying epitopes with high antigenic properties from each antigen, the epitopes were linked using an appropriate linker (EAAAK). The physicochemical properties and antigenicity of the designed antigen were then assessed. Subsequently, the three-dimensional (3D) structure was predicted and validated. The amino acid sequence of the multiepitope antigen was converted to a nucleotide sequence, optimized, and inserted into the pET23a (+) vector. Results: Bioinformatic analyses indicated that the designed multi-epitope antigen is stable, soluble, and possesses significant antigenicity. Conclusion: Our findings suggest that the designed multi-epitope antigen could be a promising candidate for the diagnosis of brucellosis. However, further validation through laboratory methods is necessary to confirm its diagnostic efficacy. [ABSTRACT FROM AUTHOR]

Background and Objectives Umbilical cord blood is a valuable source of stem cells used in transplants to treat various diseases including leukemia, lymphoma and genetic disorders. However, cord blood clotting during the collection process can reduce sample quality and quantity and impact its efficacy in cord blood banking. This article aims to predict pre-collection cord blood clotting in donors using advanced machine learning techniques. Materials and Methods In this retrospective study, data was gathered using 928127 samples available in the fetal cord blood bank, and with using supervised machine learning classification algorithms, including decision tree, naïve Bayes, K-Nearest Neighbors, Support vector machine, Random forest, Majority voting and Multilayer perceptron, prediction of cord blood clotting was performed on the Royan cord blood bank database and their performance was compared using evaluation metrics such as Accuracy, Precision, Recall, and F1 Score. Results In this study, the algorithm accuracy of Decision Tree was 0.80, Naive Bayes was 0.63, KNearest Neighbors was 0.83, Support Vector Machine was 0.65, Random Forest was 0.84, Majority Voting Classifier was 0.81, and Multilayer Perceptron was 0.74. Conclusions In this study, the performance of Random Forest and K-Nearest Neighbors algorithms demonstrated the best accuracy showing that machine learning algorithms can predict prenatal cord blood clotting with high accuracy which can help prevent sampling of clotted specimens in order to reduce costs and storage problems. [ABSTRACT FROM AUTHOR]

Objective: Mastitis, a significant infectious ailment affecting dairy cows during both dry and lactation periods, has substantial impacts on animal health, economic profitability, and the dairy industry each year. This study seeks to utilize bioinformatics analysis not only to identify hub genes and biological pathways linked to mastitis in dairy cows but also to gain a deeper understanding of its implications. Materials and methods: 21 microarray data samples were obtained from the GEO database (accession number GSE24217), comprising 9 healthy cow tissue samples and 12 E. coli infected tissue samples. These samples were assessed using GEO2R. Differentially expressed genes were identified based on two criteria: adjP-value > 0.05 and |LogFC| ≥ 1. Hub genes were identified using the CytoHubba plugin in Cytoscape software and topological analysis methods (DMNC, MCC, MNC, DEGREE). Signaling pathways were identified using the STRING tool based on an FDR > 0.05 index. Results: Out of 631 differentially expressed genes, 136 had low expression, and 495 had high expression in infected breast tissue compared to healthy ones. Among these, 205 genes were co-expressed in the network. 12 hub genes, including CCL19, ALB, GAPDH, PTPRC, ICAM1, IL6, IL1B, IL18, CXCL8, CCL20, CXCL16, and CCL3, were identified. The analysis of biological pathways revealed that the studied genes are involved in 66 biological pathways, with NOD-like receptor signaling pathway, TNF signaling pathway, and IL-17 signaling pathway playing important functional roles in dairy cow mastitis disease. Conclusions: The discovered genes and pathways have the potential to enhance our knowledge of the complex molecular mechanisms involved in mastitis development. These discoveries could shape a thorough reform initiative focused on preventing, diagnosing, and treating mastitis. These insights are expected to pave the way for the creation of precise and dependable biomarkers for early detection and prevention of bovine E. coli mastitis, ultimately facilitating personalized treatment approaches. [ABSTRACT FROM AUTHOR]

Introduction: Since the introduction of Next-Generation Sequencing (NGS) in the early 2000s, this technology has emerged as a transformative advancement in the life sciences, significantly propelling genomic, transcriptomic, epigenomic, and other related research fields. The core principles of NGS technology encompass library preparation, sequencing, and the analysis of the resulting data. By enabling the parallel sequencing of millions of DNA fragments with high accuracy, low cost, and rapid turnaround, NGS has effectively replaced older methods like Sanger sequencing. It has revolutionized our understanding of genetic complexities, genome structures, and genetic diversity through the swift and precise sequencing of entire genomes and target regions. Key applications of NGS in the life sciences include the identification and study of genes related to quantitative and qualitative traits, genetic diversity studies, population genetics, the diagnosis of genetic diseases, epidemiology, microbiome analysis, forensic science, phylogenetics, systems biology, genetic engineering, genome editing, and plant and animal breeding. However, the effective use of NGS data necessitates the development of robust computational infrastructure and advanced algorithms, as well as the expansion of researchers' knowledge regarding the bioinformatic applications and challenges associated with NGS data analysis and interpretation. Materials and methods: The present article is a review paper, conducted through content analysis by searching for keywords related to Next-Generation Sequencing (NGS), types of NGS sequencing, NGS data analysis, and the applications of NGS in relevant articles found in online databases such as PubMed, Web of Science, Google Scholar, and Scopus. Results: This study aims to provide a comprehensive guide for the efficient and optimal analysis of NGS data by thoroughly reviewing first-, second-, and third-generation sequencing methods, examining NGS data analysis pipelines, and exploring the broad applications of NGS in various fields, including cereal research. The first section reviews first-generation sequencing (Maxam-Gilbert and Sanger), second-generation sequencing (Illumina, ABI/SOLID, Roche/454 pyrosequencing, Ion Torrent), and third-generation sequencing (Heliscope, SMRT, and Oxford Nanopore). The second section introduces various NGS sequencing methods, such as Whole Genome Sequencing (WGS), Whole Exome Sequencing (WES), Bulk RNA-Seq, and others, and examines their analysis pathways. The subsequent discussion elaborates on the application of NGS in diverse areas, including the identification of structural genomic variations (SVs), the study of epigenetic changes, microbial population analysis, and agriculture (with an emphasis on cereal breeding). Finally, the advantages and challenges of NGS are discussed. Conclusion: As a revolutionary technology in genomics, Next-Generation Sequencing has profoundly impacted life sciences research. The reduction in sequencing costs, coupled with increased accuracy and the development of new methods, has positioned NGS as a critical tool for a deeper understanding of genetics and the development of personalized therapeutic strategies. With ongoing advancements in this field and the integration of NGS with artificial intelligence, the future of NGS in enhancing the precision of genetic data analysis and improving therapeutic processes appears promising. [ABSTRACT FROM AUTHOR]

IntroductionPotato (Solanum tuberosum L.) is an economic crop that is grown and consumed worldwide. Potato is considered as the fourth important crop for human consumption after maize, wheat, and rice. Potato contain more carbohydrates, proteins, minerals, and vitamins per unit area than cereals. However, this crops exposed environmental stresses during growth and developmental stages particularly, drought stress. Potato is sensitive to drought condition because of narrow root system. As a result, it is necessary the investigation of molecular mechanism involved in drought stress in potato. Drought stress affected on the integrity and structure maintain of biological membrane. The actin-based myosin is necessary for the growth and organization of the endomembrane system. Given that myosin genes allow to move intercellular and intracellular providing the possibility of repair of damaged membrane areas under drought stress in plants. In eukaryotic cells, organelle movement, positioning and communications are critical for maintaining cellular functions and are highly regulated by intracellular trafficking. Directional movement of motor proteins along the cytoskeleton is one of the key regulators of such trafficking. Most plants have developed a unique actin–myosin system for intracellular trafficking. Myosins participate in a different cellular processes, endocytosis, containing cell polarization, intracellular and organellar transport, transcriptional regulation, and signal transduction. Motions are as molecular motors in biological systems. These can bind to filamentous actin and produce physical forces through hydrolyzing ATP. Myosins is one of the most diverse and largest protein families in eukaryote. Class XI and VIII motor proteins are characterized as a slow and fast motor proteins. Myosin XI possess many genes in Angiosperms. Whereas, myosin-VIII contain fewer members against class XI. Further, molecular size of myosin-VIII is smaller than myosin-XI.Materials and methodsProtein file of A.thaliana was downloaded and matrix file of hidden Markov Model (HMM) of myosin gene family was used. Here, phylogenetic analysis, gene structure, and gene expression were surveyed. Gene expression of two genes (StMyoXI-F and StMyoXI-B) was performed in different tissues (root, stem, leaves, and tuber). For analysis of expression in different tissues (leaves, stem, and root), sampling was performed 2 weeks after mother tuber growth. However, tubers were sampled 6 weeks after cultivated in Karaj. To evaluate expression under drought stress, after 6 weeks of growth, the water stress was induced by irrigation suppression in 2 weeks while, daily irrigation was maintained for control plants. Leaves and tuber sampling were performed under two conditions.Results and discussionThe seven StMyo genes were unevenly distributed in the two subgroups. The seven StMyo genes were unevenly distributed in the two subgroups. Class XI was the largest subfamily, which class VIII had the lowest subfamily. Drought treatment significantly increased expression of StMyoXI-B and StMyoXI-F genes by 80 and 8 fold compared to the control treatment, repectively. Under control conditions, StMyoXI-B and StMyoXI-F enhanced expression in root by 25 and 20 fold while, it showed low expression in leaves under control conditions. Under control conditions, StMyoXI-F gene showed the 3 and 4 fold change higher mean relative expression than the reference gene. The relative expression of StMyoXI-B gene has been increased 5 and 10 fold change in leaves and stem under non-stress conditions as compared to the reference gene. Among myosin genes, the number of exons varied from 1 to 39. Most of class-VIII proteins had fewer introns whereas, class-XI proteins possess more proteins than class VIII. This results indicated that occurred splicing process in the myosin genes.ConclusionBased on the obtained results, it is considered that these genes highly do not influenced in stem development. However, it had significantly affected in root, leaves, and tuber development. It seems this gene can be potentially a good candidate for drought tolerance breeding programs in potato.

Introduction: In recent years, highly pathogenicity avian influenza (HPAI), especially H5N1, has emerged as a major global health concern due to its potential as a zoonotic disease and its devastating impact on poultry populations. Identifying the molecular mechanisms of response to HPAI infection is critical to control, treat, and prevent the risk of a potential pandemic. Microarray technology is becoming a standard technology used in research laboratories all across the world and it is considered as one of the centers of research in cellular processes related to the level and manner of gene expression, including gene function and cell differentiation mechanisms. By using microarray technology, it is possible to obtain a detailed view of the interaction function of genes while simultaneously studying how the genome is expressed. Using microarrays provides the analysis of gene expression in response to viral infections such as influenza, etc., the study of host-pathogen interactions, and also the identification of the effectiveness of drugs and vaccines. This study aimed to analyze the microarray data of H5N1 avian influenza to compare the gene network and analyze the functional pathway in chickens and ducks. Materials and methods: Data mining and searching of microarray data related to Highly Pathogenic Avian Influenza infection was done in the GEO gene expression database (https://www.ncbi.nlm.nih.gov/geo). The microarray data set with accession number GSE33389 based on the GPL3213 platform was selected which contained lung tissue samples challenged with H5N1 virus in chickens and ducks. Normalization of selected microarray data was done using R software, and samples were grouped to compare between infected and control samples. Limma, Biobase, and GEOquery software packages in R software were used to determine the expression level of genes and to investigate the differentially expressed genes (DEGs) between healthy and H5N1 influenza virus-infected lung tissue samples in chickens and ducks. The criterion for selecting significant DEGs was considered as |logFC|>2 and P<0.05. DAVID online tool (https://david.ncifcrf.gov) was used to investigate biological pathways, structural and functional characteristics of genes with different expressions, and functional interpretation of upregulated and downregulated DEGs. It was evaluated and visualized separately based on biological processes (BP), molecular functions (MF), and cellular components (CC). KEGG tool (http://www.genome.jp/kegg) was used to evaluate and study metabolic pathway enrichment. To reveal interactions between proteins and analyze them, STRING database and Cytoscape software were used. While using the Cytohubba plugin to identify and display key genes, the main modules affecting the interaction of genes and proteins were also identified by the MCODE plugin. Results and discussion: Gene expression analysis revealed 2062 and 565 differentially expressed genes between normal and infected tissue in chickens and ducks, respectively (P<0.05 and |logFC|>2). The results of bioinformatics analysis and protein-protein interaction network analysis showed BUB1, NDC80, CDC20, PLK1, PRC1, KIF11, and AURKA genes as hub genes in chicken and also COL6A3, COL3A1, COL4A3, COL18A1,PLOD2, PLOD1, and P4HA2 as highly effective genes in duck (P< 0.05). The results of the ontology comparison of DEGs proved that most of these genes in chickens are involved in the innate immune response and inflammatory resistance of the host, and the most effective genes in ducks play a role in lipid metabolism and energy production to meet the host's resistance to disease. The results of comparative gene network analysis between chickens and ducks are promising to increase our understanding of the host response to H5N1 influenza infection and the factors affecting virus pathogenesis in different avian species. Differentially expressed genes in response to H5N1 infection in chickens and ducks play critical roles in various biological processes, including immune response, inflammation, viral replication, and host-pathogen interactions. In general, gene network analysis showed that chickens and ducks use different genetic strategies to respond to avian influenza virus infection. Conclusions: The present research was conducted to discover the response to H5N1 HPAI infection in chickens and ducks through comparative gene network analysis. It is important to note that the gene network analysis presented in this research is an initial step towards discovering the response mode of HPAI (H5N1) infection in chickens and ducks, and further functional studies, validation experiments, and integration with other omics data are needed to confirm the role of genes, pathways and hub genes in the host response to H5N1 virus. Therefore, the results of comparative gene network analysis in chickens and ducks obtained from this research can provide valuable insight into the underlying molecular mechanisms of host response to H5N1 influenza infection. Thus, by identifying differentially expressed genes, functional modules, and hub genes in this research, it can be stated that potential targets for future research have been highlighted to some extent. Undoubtedly, further studies in this field will improve our knowledge about the pathogenesis of avian influenza and will help to develop strategies for effective control and prevention of H5N1 influenza outbreaks. [ABSTRACT FROM AUTHOR]

IntroductionSoil and water salinity has significantly reduced crop yields and threatened human food security worldwide. Understanding the molecular basis of how crop plants receive and respond to salinity and identifying the main components of their stress tolerance are essential for developing tolerant plants through genetic manipulation. Today, functional genomics methods such as transcriptomics, proteomics, and metabolomics have revolutionised the study of plant stress response and facilitated the identification of important pathways and components governing stress tolerance. For example, the production and analysis of EST sequences, RNA microarray, and RNA-sequencing technology (RNA-seq) have made it possible to study and identify gene transcripts in genome-wide stress response networks. Accordingly, this study aimed to analyse the transcript of forage turnip (Brassica rapa L.) under salinity stress, determine the functional orientation of the genome, and identify the important components and genes involved in salinity tolerance.Materials and methodsTwo Expressed Sequenced Tags (ESTs) libraries of forage turnip under non-stress and salinity stress conditions were analysed using bioinformatic and statistical methods. Non-stress and salinity stress libraries had 8408 and 6894 sequences, respectively. After sequences trimming, the genome functional orientation of forage turnip in response to salinity stress was determined based on Fisher's exact test using the DAVID tool. Differentially expressed genes (DEGs) were assigned using the Audio and Claverie statistical test (AC test) implemented in the IDEG6 tool, at a significance level of 5%. Based on Fisher's exact test, a hierarchical gene network among DEGs was predicted. Then, gene network topology analysis was performed using the NetworkAnalyzer plugin in Cytoscape 4.3. One gene was identified as an important gene (Hub gene) based on gene network topology analysis. In a greenhouse experiment using a tolerant genotype and a susceptible genotype of forage turnip, the expression profile of the identified important gene along with some traits related to the plant antioxidant system, including anthocyanin content, antioxidant enzymes activity, and malondialdehyde content, were evaluated in response to different salinity time points. Finally, the relationship among the changes in gene expression, evaluated traits, and salinity tolerance was determined using Correspondence Analysis (CA).Results and discussionPre-processing of 8408 EST sequences of the non-stress library resulted in 8,403 high-quality sequences. Also, out of 6894 EST sequences from the salinity stress library, 6894 high-quality sequences were obtained. BlastX against Arabidopsis proteins showed that 8075 (96.1%) of the non-stress EST sequences had at least one specific hit. 6597 (95.7%) of the EST sequences of the salinity stress library also had at least one specific hit in BlastX. Based on Fisher's exact test, 15 functional groups were significantly (FDR≤0.01) more active in salinity stress conditions than in non-stress conditions. These results clearly showed that under salinity conditions, the genome functional activity of forage turnip was oriented towards response to stresses, especially oxidative stresses, response to internal inductions such as plant hormones, control of metabolic activities, and photosynthetic reactions. Audic and Claverie’s statistical test showed 344 genes were differentially expressed in response to salinity stress; among them, 242 genes were upregulated, and 102 genes were downregulated. The predicted gene network indicated a complex relationship among DEGs, regulatory molecules (especially melatonin and plant hormones), and downstream responsive pathways. Among identified DEGs, the gene encoding transcription factor ZAT6 was assigned as an important gene in the salinity response gene network. The expression profile of the ZAT6 gene, quantity of anthocyanin, the activity of antioxidant enzymes and content of malondialdehyde were significantly different between the two studied genotypes. ZAT6 was significantly more expressed in the stress-tolerant genotype than the stress-susceptible genotype at all time points. In addition, the antioxidant system of the tolerant genotype was more potent than the susceptible genotype. Also, results revealed a significant relationship between the expression profile of ZAT6 and evaluated traits in the context of salinity tolerance. Based on the results, changes in ZAT6 gene expression are directly or indirectly involved in regulating forage turnip plant responses to salinity stress, especially through the control of evaluated traits.ConclusionTranscriptome study clarified some of the molecular bases of the forage turnip response to salinity stress. Accordingly, it seems that the gene encoding the ZAT6 transcription factor plays an important role(s) in the salinity stress tolerance of forage turnip. There was a significant relationship between high expression levels of this gene and enhanced antioxidant activities, which could confirm the hypothesis. However, further studies are needed to assign detailed functions of ZAT6, particularly the association of this gene with the regulatory pathway of melatonin as a major regulatory molecule in plants. This can be an effective starting point for further studies.AcknowledgmentsWe would like to thank the support of the Higher Education Center of Eghlid for this research project.

Introduction: Stomach cancer is the fourth most common cancer and the second leading cause of cancer death in the world. In addition to significant costs, it imposes many problems on families that is a proper justification for identifying related biomarkers. Method: The miRBASE website was used to find bioinformatics information. In addition, mirWALK and miRNASN databases were searched to find miRNAs information. To find cancer pathways, the DAVID and KEGG databases were adopted. Results: The studies conducted on the first pathway reported that microRNA and has-miR-10a-3p, through binding to rs1049216 in the CASP3 gene, by exerting an inhibitory effect on this gene, prevents the development of cancer. In the second pathway, microRNA and has-miR-296-3p, by binding to rs1564483 in the BCL2 gene, by exerting an inhibitory effect on this gene, causes cancer. In the third pathway, microRNA and has-miR-194-3p, through binding to rs7177 in the CCND1 gene, by exerting an inhibitory effect on this gene, prevents the development of cancer. Conclusion: In this research, cancer pathways involved in cancer were predicted. By finding three SNPs on specific regions of the three above-said genes, we found that these SNPs can be the cause of stomach cancer and can be introduced as biological biomarkers.

Introduction: Large quantities of dyes, including refractory azo dyes, are discharged directly into wastewater from the textile and petroleum industries and can be removed by bioremediation. In this study, bioinformatics tools were used to modify the structure of the enzyme azuridactase to improve its performance in degrading these dyes, including methyl red. Methods: The amino acid sequence of the azoridactase enzyme was obtained from the UniProt database. The three-dimensional structure of the enzyme was predicted using modeling tools, and the best model was determined using Qmean web software. Due to the close proximity of the active site of this enzyme to that of Bacillus Smithii, the substrate (methyl red) was docked to a three-dimensional model of the active site using the PyRx program. Potential mutations at the active site were identified through sequence alignment. The exerted mutations were examined regarding the changes in binding energy and the interaction network. Results: The structure generated by Robetta was chosen as the best model for the Q9X4K2 sequence. The mutagenesis results, in terms of binding energy and interaction plot, indicated that the optimal mutation involves changing proline 132 to serine. This mutation reduces the binding energy between methyl red and azoridactase from -6.9 kcal/mol to -7.4 kcal/mol. Furthermore, an examination of the interaction network in the mutant protein revealed the formation of a new hydrogen bond. Conclusion: The reduced binding energy between the enzyme and methyl red suggests that the enzyme is more favorably positioned towards the substrate, thereby enhancing the enzyme's efficacy in degrading azo dyes. [ABSTRACT FROM AUTHOR]

Introduction: Polymerase chain reaction (PCR) is a critical and practical technology in various biological and medical sciences fields. Using this technique, millions of copies of a specific pattern can be reproduced in vitro with a minimal amount of DNA. This has made significant advances in molecular biology and medicine. This technique has many applications in biological sciences. PCR primers are an integral part of PCR. Well-designed primers allow the PCR system to work accurately. To create a good PCR primer pair, several factors must be considered, including base pair length, melting and annealing temperature, base pair repeats, etc. Carefully designed PCR primers increase sensitivity and specificity and reduce the effort spent on experimental optimization. The poorly designed primer and the lack of optimization of the reaction conditions probably lead to decreased technical accuracy and false positive or negative detection in the amplification of the target DNA fragments. Bioinformatics has become an essential tool for basic research and applied research in life sciences. Primer designing tools provide optimal and efficient design of primers by automating complex and accurate calculations in the shortest possible time. Materials and methods: This review article is a content analysis study performed by searching Polymerase Chain Reaction (PCR), PCR type, and primer design for PCR in related articles on Google Scholar, Web of Science, PubMed and Scopus. Results: This article aims to provide an almost complete guide for designing efficient and optimal primers for PCR and qPCR analyses by reviewing the PCR method, its types and applications and using the links and web-based services. In this regard, in the first part of the article, various methods, such as standard PCR, reverse transcription PCR (RT-PCR), quantitative PCR (qPCR), combined RT-PCR/qPCR, etc., will be explained with suitable figures. The following summarizes the use of PCR types, emphasizing cereal research. In the end, primer design methods, especially for qPCR, classification and review of existing software and their capabilities, methods and software for testing the features of primers to obtain maximum specificity and PCR efficiency, extracting the gene sequence for PCR primer designing is presented with practical examples. Conclusion: A PCR with weak primers can produce little or no product due to non-specific amplification or primer dimer formation. Several online tools were introduced that are used to prepare effective primers for PCR in the field of molecular biology. [ABSTRACT FROM AUTHOR]

Apocarotenoids play a pivotal role in the economic value of saffron plants, underscoring the significance of isolating and studying the genes implicated in apocarotenoid metabolism. This research endeavor focused on the CsUGT gene, a key player in crocin biosynthesis. The gene was subjected to isolation, homogenization, and subsequent transfer into the E. coli strain DH5α. The gene underwent cloning, sequencing, and registration within the NCBI database. A multi-step analysis was initiated to gain a deeper understanding of the attributes inherent to this gene. Firstly, the protein sequence of CsUGT was obtained. Subsequently, several servers and tools, including Protparam, SOPMA, ProtScale, Pfam, ProtComp, SignalP, TMHMM, and ChloroP, were employed to delve into the protein's physical, chemical, and physiological characteristics. Moreover, the Swiss-Model server was leveraged to investigate the protein's three-dimensional structure. The outcome of this exploration included the construction of a Ramachandran diagram, which effectively validated the 3D model structure produced. According to the results, CsUGT protein with 462 amino acids has the conserved sequence of glycosyltransferase family proteins and was identified as a polar protein, stable at high temperatures and without a hydrophobic domain. CsUGT protein has no peptide signal or binding signals and has a cytoplasmic location. This research made it possible to isolate the saffron's CsGTS gene and optimized its transfer conditions into the vector. In addition, the results of CsUGT protein structure analysis provide the basis for future functional studies and can provide valuable information regarding the behavior and reaction of this enzyme in the synthesis of saffron apocarotenoids; also, these results can be useful in future programs of Iranian saffron genetic modification. [ABSTRACT FROM AUTHOR]

Introduction: The importance of effective treatment for prostate cancer cannot be ignored. Maintaining the quality of life of people with this common type of cancer is vital. Identifying new treatments through modern sciences such as bioinformatics is essential and provides the possibility of personalized medical programs tailored to individual needs. In addition, these advances can help develop preventive measures that can ultimately reduce the risk of developing this type of cancer. Materials and Methods: We downloaded the GSE55945 gene expression data set from NCBI-GEO and analyzed the differentially expressed genes (DEG). The role of genes in biological processes, and the enrichment pathways of genes in the Enrichr database are displayed by the Kyoto Encyclopedia of Genes and Genomes (KEGG), Reactome, Bioplanet, etc. Also, protein-protein (PPI) networks drawn and discussed. Results: A total of 7 hub or key genes, including CAV1, MYLK, CACNA1D, CALM1, NOX4, CCK, and AOX1, were identified. Analyzes related to molecular processes showed that most genes with differential expression are involved in the "mechanism of drug metabolism". Also, the results showed that the molecular function of most genes is related to "G-protein dependent", and "inositol phosphate metabolism" processes. Conclusion: DEGS, key genes and signaling pathways identified in this study may help to understand the molecular mechanisms of prostate cancer and provide possible targets diagnosing and treating this disease. [ABSTRACT FROM AUTHOR]

Introduction: Major depressive disorder (MDD) is one of the most common mental illnesses. The results obtained from previous studies conducted in this field show that inflammatory processes increase in MDD patients. However, few studies have addressed the issue of inflammatory markers in the brains of MDD patients. The aim of this study was to investigate the expression of regulatory axis related to cytokine signaling including IL6, TNF, NRG1 and BDNF in prefrontal cortex tissue samples. Materials and Methods: In the present study, using a bioinformatics approach based on microarray data analysis, the expression of a regulatory axis related to cytokine signaling, including four genes (IL6, TNF, NRG1, and BDNF), in prefrontal cortex tissue (PFC) samples was investigated. Background correction, gene filtering and data normalization were done using different packages in R. Data quality was also evaluated using AgiMicroRna package and PCA plot.The limma package in R was used in order to identify the genes with expression changes. In addition, Pearson's correlation analysis was performed in R to check the correlation between the selected genes. Results: The obtained results showed that the expression of BDNF and NRG1 genes in the PFC tissue of MDD patients was associated with a significant decrease (P= 0.037), and on the other hand, the expression of IL6 and TNF genes was significantly increased (P< 0.001). Also, a strong negative correlation between the IL6 and BDNF (P < 0.001) genes was obtained. Conclusion: In this study, the results show that there is a complex relationship between MDD disease progression and cytokine signaling. This research provides evidence for a better understanding of the mechanisms of MDD pathogenesis. [ABSTRACT FROM AUTHOR]

Introduction: The sMMO enzymes contain three common components: a hydroxylase, a reductase, and a regulatory protein. In addition to its potential role in converting methane to methanol, the sMMO enzyme oxidizes a wide range of hydrophobic molecules. The main aim of this study is to improve the stability of the sMMO enzyme by altering its amino acid sequence using bioinformatics techniques. Methods: The ClusPro web server was used for docking the hydroxylase and reductase subunits. The interactions between amino acid subunits were identified by LigPlot software. Mutagenesis was generated by Molegro Virtual Docker software based on evolutionary trends and mCSM-PP12 server predictions. The ∆G value of wild-type enzymes and mutant variants was obtained using the PRODIGY server. Results: In the Q83L, N161H, K43M, and N101S mutants, the value of ∆G was more negative than in the wild-type MMOH2MMOB complex. The results of the LigPlot analysis showed that the hydrogen bonds present in the wild-type enzyme were preserved in the Q83L and N161H mutants and that these mutations increased the stability of the enzyme. Besides, The D29T and H37F mutants cause a more negative ∆G value than the wild-type hydroxylase-reductase complex. The results of LigPlot analysis show that these mutations form a new hydrogen bond between the reductase and hydroxylase subunits, which is highly consistent with the ∆G value and increases the stability of the enzyme. Conclusion: It is concluded that changing the non-polar amino acids on the protein surface to polar ones can significantly improve the stability of enzymes. [ABSTRACT FROM AUTHOR]

Background & Aims: Schizophrenia is a mental disorder with frequent psychosis, whose symptoms include delusions, hallucinations, disordered speech or behavior, and impaired cognitive ability. The cause of this disease becoming a debilitating disorder for patients and their families is its early onset and chronic course, which affects one percent of the population. Antipsychotic drugs are only effective in treating half of the patients, and the pathological mechanism of schizophrenia is not fully understood. In addition, these drugs improve generally positive symptoms (such as hallucinations and thought disorders that are at the core of the disease), but negative (such as social isolation) and cognitive symptoms (such as learning and attention disorders) remain untreated. On the other hand, these drugs have severe neurological and metabolic side effects and may lead to sexual dysfunction. Schizophrenia is a multifactorial disorder that involves the interaction of several genes, environmental factors, and epigenetic processes. Over the past ten years, genetic research has yielded new findings that have given rise to hope for the biological origins of schizophrenia. The genetic role is caused by a large number of alleles, including alleles with small effects that can be identified by GWAS studies. The drug treatments that are available for schizophrenia have poor efficacy for many patients in which the mentioned genetic factors are very effective. Identifying the causes of this disease is an important step in the field of improving the treatment for this disorder, which is estimated to be half to one third of the genetic factors influencing the development of schizophrenia by common alleles reported in GWAS studies. Thus, GWAS studies are potentially an important tool for understanding the biological process of schizophrenia. GWAS Catalog is a complete collection of all published studies at the genome level, which makes it possible to investigate and identify the cause, understand the mechanism of the disease, and identify new treatment methods in this disease. In recent years, many efforts have been made to identify biomarkers in the diagnosis, prevention and treatment of schizophrenia, each of which introduces specific genes and pathways that overlap with other mental disorders. In this study, in order to investigate important genes and biological pathways in the development of schizophrenia, the genes reported in the GWAS catalog bioinformatics database were examined so that in the future, with more studies, it can help to treat and diagnose these patients more accurately. Methods: In the present study, using the GWAS catalog database and searching for the word Schizophrenia, a list of 3344 SNPs was extracted from 80 research articles. In the next step, the SNPs that cause schizophrenia and other related diseases were extracted (1165 SNPs that were the result of 19 studies). Then, among the SNPs obtained in the previous steps, those with lower and equal p-values were 5*10^-8, isolated and checked using ensemble.org in the BioMart section, in order to find the closest genes. Then, the results of BioMart were sorted and collected based on the obtained genes, and the duplicates were removed (removal of duplicate genes) and finally 336 genes were obtained. (In general, for 554 SNPs, the gene was identified, and for the other 574 SNPs, the gene name was not found, which are extragenic.) In the next step, the above list was entered in BioMart and the names of the desired genes were obtained. (Part of Ensemble genes 107; NCBI gene (formerly Entrezgene) accession) which resulted in a total of 339 genes. In the last step, the genes obtained from the previous step were checked using the Enrich R site, and through this site, the pathways that were related to the studied genes were extracted and compared with the results of different articles. Results: In this study, after entering the results in Biomart, 339 genes were obtained. Also, after entering the above list in the Enrich R site, the important pathways in which these genes are active were obtained. Pathways which Adjusted-P value was below 0.05 were considered as reliable results. Conclusion: This study was conducted on schizophrenia by analyzing the data of GWAS catalog database to identify the mechanism of the disease and the pathways involved as well as the main and important genes of drug candidates. One of the important pathways in this study was Glucuronidation, which was one of the first pathways obtained from Wiki Pathway studies, which is often involved in the metabolism of substances such as drugs, pollutants, bilirubin, androgens, estrogens, mineralocorticoids, glucocorticoids, fatty acid derivatives, retinoids. and bile acids play a role. This pathway is mainly carried out in the liver, although the enzyme responsible for its catalysis. UGTs are widely expressed in the brain, but at lower levels than in the liver. Brain UGTs are inducible or inhibitable that affect drug release in the central nervous system. UGTs, cytochrome P450s, and transporters act together to influence the pharmacokinetics of drugs in the brain. After glucuronidation, many drugs have the ability to cross the blood-brain barrier. Another pathway is ascorbate metabolism. Ascorbate (vitamin C) is a vital antioxidant molecule in the brain. However, it has several other important functions, participating as a cofactor in several enzymatic reactions, including catecholamine synthesis, collagen production, and HIF-1α regulation. Ascorbate is transported to the brain and neurons through the sodium-dependent vitamin C transporter (SVCT2), which causes the accumulation of ascorbate in cells against a concentration gradient. The highest concentrations of ascorbate in the body are found in the brain and neuroendocrine tissues such as the adrenals, although the brain is the hardest organ to deplete ascorbate. Together with the regional asymmetry in the distribution of ascorbate in different regions of the brain, this shows the important role of ascorbate in the brain. The next pathway was the porphyrin metabolism pathway, which disruption in this pathway can cause neurological symptoms, neuropathy and a wide range of psychiatric symptoms in addition to specific diseases, until in 1939, Waldenstrn reported that schizophrenia in families with Acute intermittent porphyria (AIP) is very common, but no statistics were provided. Another study found that 5 of 40 AIP probands had hallucinations, suggesting a possible link between psychosis and AIP. A study in Sweden found that people with AIP were four times more likely to develop schizophrenia or bipolar disorder. In addition, first-degree relatives of these people are twice as likely to be diagnosed with schizophrenia and bipolar disorder. The other pathway was the pathway of retinoid metabolism, the first evidence linking this pathway to schizophrenia is that retinoid toxicity or deficiency frequently leads to the manifestation of symptoms that, although more severe in scope, are similar to those of schizophrenia. For example, mental disorder, mental defect, large ventricles, microcephaly and various major and minor congenital anomalies, among which cranial anomalies are prominent. Such deficits have been reported frequently among schizophrenic samples. With the increasing documentation of the prominent role of retinoids in the development of the central nervous system, the possibility of retinoid involvement in schizophrenia is strengthened. Another important pathway was drug metabolism, which drugs that are widely used today in the treatment of mental disorders, including depression, psychosis, and mood stabilizers, in many cases are associated with many side effects, and only a small number of patients respond appropriately to These drugs show Many factors, including genetic factors, are effective in the effectiveness of drugs, which are of interest today. According to the present study, it can be concluded that genetics and especially polymorphisms play a very important role in the development of schizophrenia and that these genes can be used in the diagnosis and treatment of this disease. In the future, the genes of these pathways can be studied more reliably in schizophrenia for diagnostic and therapeutic purposes. The genes obtained in this research can be suitable and new options for future studies on schizophrenia and optimization of treatment methods. [ABSTRACT FROM AUTHOR]

Background and purpose: P24 is one of the important components of retroviruses capsid. Several studies have shown the inhibitory effect of melittin protein on the function of AIDS virus. The aim of this study was to evaluate the effect of this protein on P24 in SIV and HIV viruses using bioinformatics software. Materials and methods: P24 protein sequences of SIV and HIV were obtained from a gene bank and their features were analyzed using different software programs. Also, structural similarities of P24 protein between SIV and HIV viruses were examined. Docking analysis was used to examine the interactions between melittin and P24 proteins. The stability of docking results were confirmed by molecular dynamics simulation. Results: Findings showed numerous phosphorylation, glycosylation sites, and di-sulfide bonds in P24 protein. Docking results indicated a probable interaction between P24 protein of both viruses and melittin. The results showed similarity of the third structure of P24 protein between the two viruses. Analysis of molecular dynamics showed molecular stability of the interactions between P24 and melittin proteins. Conclusion: High energy values of the interaction between melittin and P24 indicated high binding potential of the two proteins, which can determine the inhibitory effect of melittin on the function of HIV capsid. Analysis of the third structure of P24 protein in both viruses showed that they are very similar despite the high difference in protein sequences from the point of view of folding and 3D structure. Application of melittin to control SIV infections in primates can be used as a model for inhibiting HIV in humans.

Background & Aims: In today's societies, breast cancer has significantly threatened women's health (1). Successful cancer treatment can also destroy tumor germ cells (2, 3). Treatment methods based on immune checkpoint molecules are considered (4). PD-L1 overexpression suppresses the immune response to cancer cells and causes epithelial-tomesenchymal transition (EMT) and cancer stem cell (CSC) phenotypes, metastasis and chemical resistance (5). Overexpression of PD-L1 and MUC1 is observed in approximately 90 TNBCs. In this present study, we investigated the inhibitory effect of miR-143-3p on MUC1 expression as an oncogene that has the most impact on PDL-1 overexpression in BC cell lines MCF-7 (less aggressive BC), MDA-MB-231, and BT549 (metastatic BC model). The suppressive effect of miR-143-3p on the expression of this oncogene as a direct effect and PDL1 level as an indirect result in BC cell lines was investigated. Besides, the effect of miR-143-3p overexpression on BC cell lines proliferation through downregulation of candidate oncogene was investigated. Methods: First, the targeting of miR-143-3P on MUC1 3 'UTR was confirmed by using bioinformatics software and then by dual luciferase assay. The expression level of miR143-3P and MUC1 were measured in breast cancer lines compared to the normal line. After transfection of miR-143-3P into breast cancer cell lines, MUC1 and PD-L1 expression in mRNA levels were evaluated by using qRT-PCR. The miR-143-3p effects on MUC1 and PD-L1 expression and its impact on cell proliferation was investigated by using MTT assay, (mean ± SD). Results: The result of the dual luciferase assay was obtained from cloning the 3'UTR of the MUC1 gene in the psi-check2 vector and sequencing. According to the prediction analysis, the 3′-UTR of MUC1 gene has been considered as a putative miR-143-3p binding site (Target scan database). Transfecting miR-143-3p into MDA-MB-231, BT549, MCF-7 cells harboring MUC1 3′-UTR decreased the luciferase activity to to 55% ± 1/73, 57% ± 1 /15, 59 % ± 3/51% respectively, in comparison with the control (P < 0.01) (mean ± SD). he expression of miR-143-3p was significantly downregulated in BC cell lines compared to their control, and its downregulation was negatively correlated with MUC1 overexpression (r= -0.6419, p = 0.0244). Increased expression of miR-143-3P, decreased the mRNA level of MUC1 in MDA-MB231 to 1/3 ± 0/7, BT549 to 1/1 ±0/45 and in MCF7 to 0/4± 0/21, compared whit untransfected cells. MiR-143-3p introduction to BC cells downregulates PDL-1 indirectly. Overexpression of miR-143-3P reduces the MUC1expression, and the reduction of MUC1 expression reduces PD-L1 expression mRNA level in MDA-MB231 to 3/4±0.45, BT549 to 3/1±0.56, MCF7 to 0/53±0.48, compared whit un transfected cells (fig4A). miR-143-3p inhibits BC cellular proliferation. An MTT assay revealed that miR-143-3p overexpression significantly suppressed the proliferation in MCF-7 to 52% ± 1.55, BT549 to 48% ± 2.45 MDA-MB231 to 43% ± 3.36, compared whit non-transfected cells 72 h after transfection. Conclusion: Breast cancer treatment has made significant progress in recent years, and a new treatment called immunotherapy, which uses the body's immune system to treat cancer, has become popular (6). One of the symptoms of breast cancer tumors is the overexpression of MUC1. A high expression level of MUC1 is associated with poor prognosis in cancer patients (7, 8). In an animal model of breast cancer, intravenous injection of MUC1 suppresses the immune system and accelerates the death of tumorbearing mice. The serum level of MUCl expression is also associated with suppression of the immune system in patients with metastatic adenocarcinoma treated with active specific immunotherapy (8, 9). As we mentioned earlier, MUC1 can increase the expression of PD-L1 by recruiting MYC and NF -kB p65 to the PD-L1 promoter in some solid tumors, such as triple negative breast cancer (10). the mechanisms underlying PDL1 expression in MUC1-positive tumor tissue were investigated, and the potential of targeting PD-L1 to inhibit the progression of MUC1-overexpressing colon cancer in mice, Therefore, the role of MUC1 in tumor immune evasion has been confirmed and can be a basis for using PD-L1 inhibitors in MUC1-positive colon cancer (8). microRNAs can regulate the expression of immune checkpoint receptors and their ligands (11). Changes in the expression of miR-143 often play a role in tumorigenesis (12). In 2020, Y Tokumaru and his colleagues showed by studying breast cancer samples with estrogen receptors that the high expression of microRNA-143 is associated with a favorable tumor immune microenvironment and better survival in breast cancer. They reported that miR143 is a tumor suppressor microRNA that exhibits anti-tumor function by targeting KRAS signalling pathways in various malignancies. The high expression of miR-143 in cancer cells with a favorable tumor immune microenvironment regulates the positivity of anticancer immune cells. The suppression of pre-cancerous immune cells is related and leads to better survival of breast cancer patients (13). In this study, we demonstrated MUC1 was potential as a novel target of miR-143-3p, therefore, the dual-luciferase reporter assay verified that miR-143-3p directly targeted MUC1 and inhibited this transcription. We found miR-143-3p downregulated in breast cancer cell lines compared to the normal line via qRT-PCR. In the current study, the qRTPCR results illustrated an inverse relation between MUC1 mRNA expression and miR143-3p expression in breast cancer cell lines. We investigated indirect targeting of PDL1 via MUC1 targeting by miR-143-3p., be able to be prevent the cell proliferation in breast cancer cell line. Therefore we recognized, targeting MUC1 may be a novel strategy for BC therapy and prevent the progression of breast cancer and miR-143-3p as a promising candidate for miR restoration therapy in BC patients. [ABSTRACT FROM AUTHOR]

Background & Aims: Chirgwin et al. (1979) were the first authors who proposed the ribonucleic acid (RNA) isolation technique. Without denaturation of the structure of nucleic acid, this method breaks the cell membrane using guanidinium thiocyanate and 2-mercaptoethanol (β-met). The extraction is carried out using ethanol or ultracentrifuge, and cesium chloride gradient. This method is time-consuming, inefficient, and involves toxic and incompatible compounds for the user. Thus, the quality and efficiency of the process of extraction were developed over time and through conducting more studies. Chomczynski and Sacchi (1987) improved the method suggested by Chirgwin et al. and proposed a single-step extraction process called the AGPC method, and it used acid guanidinium, phenol, and chloroform. The advantage of this method is its higher purity and efficiency, plus this process of isolation does not include RNA fragmentation. This method has been developed and using silica columns together with magnetic beads resulted in a revolution in the process of isolating nucleic acids. Extraction of viral nucleic acid includes high-purity DNA and RNA, determining sensitivity, and genome viral load within a short process is quite crucial for the majority of downstream molecular and medical studies such as sequencing, real-time PCR, PCR for research or diagnostic purposes. Efficient protocols for the extraction of nucleic acids are the prerequisite of these methods. As already mentioned the most available protocols are time-consuming and have limitations regarding some samples or disparate types of nucleic acids. An optimal protocol must provide a high sensitivity concerning the extraction of the viral genome from a broad spectrum of samples together with a shorter scientific time and lower price. The majority of methods up to the present time have been using precipitation steps to obtain pure nucleic acids from the extracts. New protocols use the binding capacity of the nucleic acid extract to the silica column. Vogelstein and Gillespie were the first scholars to describe the absorption of nucleic acid to the surface of silica in the presence of great concentrations of chaotropic salts. They used pieces of DNA recovered from agarose gel using silica powder. This technology is developed over time and now selective binding of DNA or RNA through silica gel surfaces is modified and binding and wash buffers have been optimized to provide the maximum separation and isolation capacity in nucleic acids. After the initial lysis of the sample, the respective nucleic acid bound to the membrane is regulated. Polysaccharides and proteins in the extract fail to function properly and lose their ability to bind to the membrane due to the destruction of the molecular structure. Finally, nucleic acid bounded to the column is desalinated by alcohol and washed, and then, separated from the column by elution buffer. One of the limitations of the process of extraction is separating various types of nucleic acids from different viral resources. For instance, Coronavirus contains an RNA genome and on the other hand, the samples are collected from the patient’s nose and throat, which requires an efficient and extremely sensitive procedure for extracting the genome of this virus. Given that, Tehran Cavosh Clon (former Sinaclon), a knowledge-based complex, proposed an optimal method for spearing viral RNA with high efficiency and free from toxic materials in accordance with conducted studies. The present research reported the results of the clinical studies on the basis of European Pharmacopoeia standards. Methods: Viral RNA was extracted from HCV, HIV, and COVID-19 samples. Sensitivity, specificity, verification, and stability tests were carried out according to the European pharmacopeia standard. Furthermore, Tehran Cavosh Clon’s viral extraction kit (SinaPure Viral) was compared with extraction kits of QIA Gen and GeneAll companies, as valid and the most common brands in the market. Finally, the results of PCR and Real-Time tests by these kits were compared. Serums infected with HCV, HIV, and COVID-19 swab nasopharyngeal were provided from Keyvan Laboratory. The reference viral genome extraction kits (QIA Gen:CAT:52904; GeneAll: CAT:128-150), and HCV, HIV, and COVID-19 diagnostic kits (GenproofHIV1/ISEX/050:HCVD/ISEX/050; CMV/ISEX/050; Sansure Biotech: S3104E) Were purchased from the respective agencies. Chemicals used in the kit structure were purchased from Sigma Company. Kit comprises lysis buffer, precipitation buffer, wash buffer I, wash buffer II, and elution buffer, carrier RNA, and binding columns. First, 400 μl lysis buffer together with 5-6 μl of carrier RNA were added to microtubes containing the samples and they were vortexed for 20 seconds. Then, 300 μl of the precipitation buffer was added to the above solution and vortexed for 5 seconds. The final solution was transferred to the column and centrifuged at 12,100 x g,13,000 rpm, for 1 minute. It must be noted that the transferred solution must be discarded. In the next step, 400 μl of wash buffer I was poured on the column, and centrifuged, at the aforesaid revolutions for 1 minute. The transferred solution was discarded. 400 μl of wash buffer II was transferred to the above column, and centrifuged, at the aforesaid revolutions for 1 minute. The transferred solution was discarded. Finally, without adding any solution, the collector column was centrifuged for 2 minutes. Then, it was transferred to 1.5 ml sterile microtubes and 50 μl elution buffer was added to it. It was placed at 55 degrees centigrade for 3 to 5 minutes. Upon the final centrifuge, the viral RNA was separated from the column with elution buffer. The extracted nucleic acid was collected. This product is used for diagnostic tests. The data were statistically analyzed using SPSS, 18. The diagrams were illustrated using Excel 2010. The One-way ANOVA test was carried out. The means were compared using Duncan's test with a significance level of P<0.05. Results: Table 1 shows the results of comparing the sensitivity test of SinaPure viral extraction kit with the QIA Gen and GeneAll regarding samples with HIV, HCV, and COVID-19. According to these results, the obtained Ct for LOD of HIV samples indicated no significant difference between GeneAll and SinaPure viral kits. While there was a significant difference between the QIA Gen and the aforesaid kits. The results of the sensitivity test of samples with HCV and COVID-19 manifested a similar pattern. The only significant difference between the QIA Gen and SinaPure viral with respect to the amount of LOD. Table 1 shows the results of the specificity of the SinaPure viral kit on different viral, bacterial, plant, and animal samples. Per the results, except for viruses with RNA such as HCV, HIV, and COVID-19, no positive response was observed in none of the viruses with DNA such as HSV, EPV, HPV, CMV, HBV, plus gram-negative bacteria, (Ecoli), gram-positive bacteria (Staphylococcus aureus), plant samples (wheat), and animal sample (CHO Cell). This confirms SinaPure Viral’s ability in extracting the genome of viruses with RNA. SinaPure Viral confirmation kit was compared to QIA Gen and GeneAll kits in high, medium, and law counts of HCV and HIV serums, plus Swab Nasopharyngeal for COVID-19. The results of disparate counts of HIV-positive samples using all three kits demonstrated no significant difference (Table 2). On the other hand, the same pattern was observed in HCV samples (Table 3). Table 4 shows the results for the COVID-19 samples. Even though kits manifested no significant difference in the high and medium counts, in lower counts of the virus, QIA Gen had a significant difference with SinaPure Viral and GeneAll. It can be due to the difference in the sensitivity of sampling since in the lower loads of virus there is a possibility of making mistakes by the sample collector. On the other hand, the lack of significant difference between the HIV and HCV samples in disparate counts confirms this. Tables 5, 6, and 7 show the results of investigating the stability of the SinaPure Viral kit in the time intervals of 6, 12, and 18 months. Per these results, SinaPure Viral kit manifested no significant difference with QIA Gen and GeneAll in the time intervals of 6 and 12 months. Even after after18 months, the SinaPure Viral kit manifested a significant difference with reference kits in low viral counts in all samples. These high count viruses did not demonstrate this reduction of efficiency. The possible cause of reduction of kit efficiency could be the location of storing carrier RNA after the first time they were used. Taking into account the considerable role of carrier RNA in the viral genome precipitation, after preparing the carriers RNA solution, its storage condition is extremely effective on the stability of RNA. Conclusion: In accordance with the results, the pharmacopoeia standard of the kit designed by Tehran Cavosh Clon corresponded to reference kits, i.e. QIA Gen and GeneAll. Given that, this kit is comparable to reference kits and it can be considered as a valid replacement for foreign kits in the market. [ABSTRACT FROM AUTHOR]

Introduction: Abortion is a common complication that refers to the early termination of pregnancy with the death of the fetus before the 20th week of pregnancy. Previous studies show that many genes are involved in this disease, including the CX3CR1 gene, which is one of the inflammatory response genes in the immune system. The pathogenicity of these variants was determined in this study using bioinformatics analysis. Material & Methods: In this study, the effects of rs3732378 and rs3732379 mutation were predicted using bioinformatics tools including SIFT, PolyPhen2, PROVEAN, Predict SNP, and Exome Variant Server. Changes in the stability of mutant proteins were investigated using I-Mutant and DynaMut tools. Moreover, modeling of the protein structure, docking, and protein-ligand interaction were performed using SWISS-MODEL, SwissDock, and FRODOCK tools as well as PyMOL, Hawkdock, and MolSoft software, respectively. Findings: Many polymorphisms related to the CX3CR1 gene have been known to date. Out of 244 missense mutations in the dbSNP database, two variants (rs3732378 and rs3732379) have been reported in association with recurrent pregnancy loss related to the CX3CR1 gene. The results of bioinformatics analyses showed that both variants were predicted as pathogenic mutations and changed the stability of the protein structure and played a key role in interaction with the ligand. Discussion & Conclusion: The findings of this study indicate that two missense mutations in the CX3CR1 gene are an important candidate for recurrent miscarriage and their identification in patients with recurrent miscarriage can be regarded as a risk factor.

Background & aim: Over the past 15 years, significant insights have been gained into the roles of miRNAs in cancer. In various cancers, miRNAs can act as oncogenes, tumor suppressors, or control the metastasis process by modulating the expression of numerous target genes. The aim of the present study was to identify molecular network of miRNA-mRNA regulating lung cancer invasion, by bioinformatics approaches. Methods: In this experimental study that was done in 2022, gene expression profiles of patients with lung cancer were collected from the RNASeq data of Cancer Genome Atlas (TCGA), by TCGAbiolinks package. Differentially expressed miRNAs and mRNAs were identified using “limma” and “edgeR” R packages and their targets were predicted by 2 databases; miRWalk, and Targetscan. Subsequently, the interaction regulatory network of miRNA-mRNA visualized using Cytoscape software. Data analysis was performed using Testing Multiple Hypotheses and False Discovery Rate. Results: The results of bioinformatics analysis of the lung cancer invasion specific miRNA-mRNA regulatory network, showed that six hub miRNAs including; hsa-let-7c-5p, hsa-let-7b-5p, hsa-let-7e-5p, hsa-miR-6838-5p, hsa-miR-320b and hsa-miR-4458, are regulator of the lung cancer invasion up-regulated genes and four hub miRNAs are regulator of this situation down-regulated gene including; hsa-miR-92a-3p, hsa-miR-204-5p ,hsa-miR-24-3p and hsa-miR-506-3P. Conclusion: The results in our study suggest that a specific miRNA-mRNA network is associated with the lung cancer invasion which this new insight into the molecular mechanism of lung cancer invasion result in the identification of molecular biomarkers and therapeutic targets for this malignancy.

Background: The heparin-binding hemagglutinin (HBHA) protein belonging to Mycobacterium tuberculosis is known as a molecular adjuvant. Objective: Hence, the expression of this protein in the prokaryotic system is essential. Methods: To predict an appropriate signal peptide for the expression of the HBHA protein, 50 signal peptides were selected from the signal peptide database. Then, the crucial parameters of signal peptides, including the probability of signal peptide, different regions of signal peptides, physicochemical features, sorting of signal peptides, and sub-cellular location were completely investigated by reliable tools. After the best-predicted signal peptide was identified, it was linked to the HBHA protein, and its secondary structure, tertiary structure, and in silico cloning in pET21a (+) was assessed. Findings: The results of different evaluations confirmed that only 13 signal peptides passed all features, including clearance of N, H, and C regions, D-score >0.7, instability index >40, and periplasmic localization. Finally, based on D-scores, among these 13 signal peptides, the asr (acid shock RNA) peptide with D-score=90 was selected as the best-predicted signal peptide to apply. Moreover, the results showed that the secondary structure of the adjuvant linked to asr peptide contained 88.18% alpha helix and 9.5% random coil. Also, the results of in silico cloning showed that the nucleotide sequences of the adjuvant linked to the asr peptide were successfully cloned between BamHI and XhoI enzymes in the multiple cloning site of pET21a (+). Conclusion: The results of this study confirmed that the asr peptide can be used as an appropriate signal peptide for the expression of the HBHA protein in the prokaryotic system.

Background and Objectives: Acute lymphocytic leukemia (ALL) is a type of cancer that affects white blood cells. The disease progresses rapidly mandating immediate treatment. ALL is the most common type of leukemia affecting children. An increase in the miR-222-3p molecule has been observed in the course of this disease. The microarray method can be used to examine the binding of this miRNA and target genes, which is not without limitations. Hence, the present study deployed computer and bioinformatics methods with different specialized software. Materials and Methods: Target genes were identified using different algorithms available in Mirbase, TargetScan, NCBI and RNAhybrid databases, and based on the scores obtained, the targets with the highest affinity were identified. Results: The findings of this study showed that the CDH4, AJAP1, HNRNPA0, UBN1 and ZNF618 genes are potential targets with higher specificity for miR-222-3p. Conclusions: Factors involved in selecting targets with high specificity and affinity is dependent on the amount of negative free energy and the number of base pairs formed between miRNA and 3'-UTR of different genes.

miRNAs genes are regulatory elements that their main role is to downregulate gene expression at the mRNAs level. miRNAs also play important roles in several plant pathways related to important cellular activities such as growth, reproduction, differentiation, morphogenesis, apoptosis and, response to abiotic and biotic stresses. This research was conducted to identify conserved miRNAs and their target genes using next generation sequencing data in the damask rose (Rosa damascena Mill.). Although bioinformatics methods have been developed as the most efficient strategy for target miRNA identification, high-throughput experimental strategies are still in high demand. Bioinformatics tools (Cmii, psRNATarget, WEGO, Blast2GO) and laboratory method (real-time PCR) were used to identify new miRNAs, predict and determine the ontology of the target genes involved in the production of scent and color in the damask rose, respectively. Using EST and RNA-seq data based on Rosa lucieae homology, four miRNA families, including: miR5021, miR2673, miR156, and miR838, were selected as candidate miRNAs. In the next step, for the quantitative evaluation of the real-time PCR to validate the expression level of the selected miR5021 in two samples (white and hot-pink) and the biological stage (young and bud) of the damask rose, foliar spraying with the concentrations of zero (control) and 300 μM of methyl Jasmonate for 48 hours was made. Result showed the highest relative expression of miR5021 belonging to the pink sample on the young development stage. These findings accelerate future prospective studies on the regulatory mechanisms of miRNAs in Rosa damascena. [ABSTRACT FROM AUTHOR]

Introduction: Long non-coding RNAs (lncRNAs) and miRNAs belong to a class of non-coding RNAs (ncRNAs) that play important roles and functions in the regulation of the expression of genes in main biological processes, such as cell proliferation, apoptosis, and differentiation. LncRNAs can potentially affect miRNAs in the forms of cis/trans to modulate their regulatory role. In this study, mRNA, miRNA, and lncRNA gene networks were predicted by web-based programs for three ectodermal pathway markers (BMP4, NOG, FGF8) in the mouse embryonic stem cells. Material & Methods: In this theoretical bioinformatics study, the miRNAs of the target genes (BMP4, NOG, and FGF8) were extracted and examined by MirWalk and TARGETSCAN databases to finally obtain the common miRNAs of these three genes. Following that, the target lncRNAs for common miRNAs were then extracted from the DIANA-Tool database. (Ethic code: 100/21560/2/پ) Findings: MiRs mmu-miR-92a-2-5p, mmu-miR-129b-5p, mmu-miR-130b-5p, mmu-miR-692, mmu-miR-7009-3P, mmu-miR-7116-3p, and mmu-miR-7689-3p may affect the function of lncRNAs, including Kcnq1ot1, Gm26812, Gm4117, Gm11837, 4930423MO2Rik, Malat1, Gm12594, Gm3414, 5830444B04Rik, Gm2464, and NEAT1. Discussion & Conclusion: Due to the mutual relationships among lncRNA, miRNA, and mRNA, our results provided a novel perspective on lncRNAs for future research and experimental studies on ectodermal differentiation pathways and molecular mechanisms.

Background and purpose: P24 is one of the important components of retroviruses capsid. Several studies have shown the inhibitory effect of melittin protein on the function of AIDS virus. The aim of this study was to evaluate the effect of this protein on P24 in SIV and HIV viruses using bioinformatics software. Materials and methods: P24 protein sequences of SIV and HIV were obtained from a gene bank and their features were analyzed using different software programs. Also, structural similarities of P24 protein between SIV and HIV viruses were examined. Docking analysis was used to examine the interactions between melittin and P24 proteins. The stability of docking results were confirmed by molecular dynamics simulation. Results: Findings showed numerous phosphorylation, glycosylation sites, and di-sulfide bonds in P24 protein. Docking results indicated a probable interaction between P24 protein of both viruses and melittin. The results showed similarity of the third structure of P24 protein between the two viruses. Analysis of molecular dynamics showed molecular stability of the interactions between P24 and melittin proteins. Conclusion: High energy values of the interaction between melittin and P24 indicated high binding potential of the two proteins, which can determine the inhibitory effect of melittin on the function of HIV capsid. Analysis of the third structure of P24 protein in both viruses showed that they are very similar despite the high difference in protein sequences from the point of view of folding and 3D structure. Application of melittin to control SIV infections in primates can be used as a model for inhibiting HIV in humans. [ABSTRACT FROM AUTHOR]

Introduction: Renal cell carcinoma (RCC) is the most common type of kidney cancer in adults. The aim of the present study was to obtain specific genes in RCC, based on bioinformatics studies and to evaluate the expression change of candidate genes in ACHN cells after transfection of the seh gene of Staphylococcus aureus. Methods: The expression correlations of genes involved in RCC were investigated in the TCGA database, and networks of genes was constructed. In vivo studies were performed in the ACHN cell model, in which the staphylococcal seh gene was cloned into the pcDNA3.1 (+) vector and transfected into ACHN cells. After confirming the correct function of the recombinant vector in these cells and inducing apoptosis, the expression of candidate genes was evaluated. Results: The results showed the successful identification of the expression correlations of genes involved in apoptosis and lncRNA-mRNA interaction network in RCC. Induction of apoptosis under the influence of toxin expression was confirmed by flow cytometry. Expression of SOX9, SBF2/AS1, MAF and c-Maf genes in cells ordered by recombinant vector was significantly reduced compared to control cells. Also, increased expression of LINC00671 and LINC02499 genes was observed in cells transfected with pcDNA3.1 (+)- seh vector compared to the control group (p <0.05) and This change in expression confirmed the accurate identification of genes involved in RCC cell apoptosis. Conclusion: The gene networks identified in this study can provide a new perspective for research aimed at identifying new molecular pathways and treating RCC. [ABSTRACT FROM AUTHOR]

Introduction Obviously, the recent decades strategy in cancer therapy, anticancer drug discovery and drug improvement is to characterize, distinguish and validate the most promising cancer-related molecular targets to which new drugs can be designed. The unique features of the pancreatic RNase (RNase A) such as high activity, stability, lack of a cofactor, and small molecular size have made it the most popular enzyme in the ribonuclease family. Specifically, RNase A is involved in endonucleolytic cleavage of 3'-phosphomononucleotides and 3'- phosphooligonucleotides ending in C-P or U-P with 2', 3’- cyclic phosphate intermediates. RNase A was purified from the Bovidae family including bovine, ovine, bison, eland, goat and gnu. Although phylogenetic analyses of RNase A revealed high similarity among members of the Bovidae family, some functional mutations were also found.Several studies showed that the RNA hydrolyzing action of ribonuclease is able to induce apoptosis and cell death in cancer cells, independently. This effect could be enhanced thousands of times when ribonuclease is linked to antibodies. These enzymes show potent cytotoxic activity on cell internalization but do not show sensible immunogenicity or non-specific toxicity toward normal cells. Ovine pancreatic ribonuclease enzyme is a member of super family RNase A, it can be a good candidate as a toxin for designing new drugs. The objective of this study was to produce ovine pancreatic ribonuclease enzyme in E. coli and characterize its activity.Materials and methods All structures needed for this study were downloaded from the Protein Data Bank (PDB) website (http://www.rcsb.org/). PDB files (accession numbers: 10.2210 /pdb1YV6/pdb and 10.2210/pdb3SNF/pdb) related to natural Bovine Pancreatic RNase were selected. Gene synthesis and production of recombinant protein were conducted by using the pelB signal sequence at the beginning of the structure for periplasmic protein production. The native ovine RNase and bovine RNase A were optimized for E. coli host by GenRay codon optimization service and sent to GenRay Biotechnology (Shanghai, China) for synthesis. The target genes in pGH vector were sub-cloned in pUC19 and then cloned into the pET21b (+) vector between XbaI and HindIII sites. After transformation, E. coli cells containing recombinant pET21b (+) were cultivated in LB broth medium containing ampicillin. To extract the proteins, osmotic shock methods were applied. After that Q-sepharose chromatography was used to extract the target protein. Finally, Bradford analysis was used to determine the protein concentration. The ribonucleolytic activity of the recombinant native ovine RNase A was compared with native bovine RNase A following Tripathy et al (2013) method. To investigate the antitumor activity of recombinant proteins, HeLa cells were prepared for seeding in a 96-well flask and incubated at 37°C in 5% CO2 for 72 h.Results and discussion The production of native ovine and bovine RNase A was confirmed by SDS PAGE. Protein purification was successfully performed using osmotic shock in the Q-Sepharose column. Although our findings confirmed protein expression, no detectable proteins other than RNase A was observed in the LB medium, indicating that almost all the proteins were expressed either inside the bacterial cell or secreted into the periplasmic area. According to the Bradford analysis, the concentration of the recombinant proteins extracted was 4.78 ovine RNase A. Based on our results, it was shown that the ribonucleolytic activity of native ovine RNase A was 558±22. The results showed that RNase A exhibited resistance to pepsin degradation during the whole incubation process (22 h), in the course of which 90% of RNase A remained undamaged. To determine the cytotoxic effect of ovine RNase on the HeLa cell line, MTT assay was done following incubation with ovine RNase A. The commercial RNase A was used as control. The results showed that ovine RNase A and bovine RNase A had no cytotoxic effect on HeLa cells. When RNase treatment was done by the lipofectamine 3000 (Thermo, USA), the cytotoxicity effect was observed. Several studies have shown that some ribonucleases such as onconase, bovine seminal ribonuclease and bovine pancreatic ribonuclease have a great promise as cancer immunotherapeutic agents and cause a significant reduction in the protein synthesis of tumor cells after internalization into cytosol. Conclusion Our findings demonstrate that ovine RNase similar to bovine RNase has a great potential for use in drug design industry. We revealed that the native ovine RNase A was more stable than the native bovine RNase. In future work, we intend to fuse the engineered ovine-RNase A to dedicated recombinant antibodies for cancer therapy and investigation of engineered immuno-ribonuclease potency and cell killing effects as a fusion protein.

Background & Aims: Antibodies, well-known as immunoglobulins (Igs), are produced by B lymphocytes and specifically defend against pathogens. Igs are glycoproteins and have high diagnostic value in several diseases including infections (1). Igs are composed of light and heavy chains (2, 3). Each chain is comprised of about 110-120 amino acid residues which create immunoglobulin folds named domains (4, 5). Domains play an important role in Igs biological functions. Every domain is included of two anti-parallel β-sheets. B-sheets play an important role in epitope formation because most of the epitopes are located in B-sheets (6, 7). Epitope/ antigenic determinant is a part of antigen recognized by antibody (8). For precise detections of Igs, diagnostic tools such as Igs- epitope specific monoclonal antibodies (MAbs) are needed (11). Therefore B-sheets are valuable tools in detection of proteins epitopes. IgG is the highest Ig in serum and has vital role in defense against infections (12). For optimizing current IgG diagnostic tests, accurate determination of IgG-specific epitopes is extremely important (18). Immunoinformatic is a branch of immunology uses a large amount of computer-generated biological information to solve immunological problems and diagnosis of diseases faster and more accurate (19-21). Immunoinformatic has spread to almost all areas of immunology and has exceedingly facilitated understanding of immune responses and their role in disease and health. Accordingly immunoinformatic has provided novel research opportunities to study the molecular mechanisms of immunological processes and related diseases and thus provide more effective diagnostic and treatment strategies (24-26). As B-sheets play an important role in epitope formation (6, 7) and immunogenic epitopes are very useful tools for production of specific monoclonal antibodies against a molecule (11), we conducted this research for recognition of human IgG B-sheets with immunoinformatic method and study their compatibility with IgG epitopes. Methods: The confirmed amino acid sequence of reference human IgG was determined with PDB data bank in http://www.rcsb.org/pdb/home/home.do web site. The second structure of human IgG was obtained with Phyre2 (Protein Homology/analogy Recognition Engine V 2.0) software available on the http://www.sbg.bio.ic.ac.uk/phyre website. The location of B-sheets and epitopes in IgG was determined by the IEDB (Immune Epitope Database) at www.immuneepitope.org. Website. Results: Figures 1 and 2 show the second structure of light and heavy chains of the human IgG molecule, respectively, obtained by phyre2 software. The IgG light and heavy chains second structure consists of alpha spirals and B-sheets. Figures 3 and 4 show the location of B-sheets in light and heavy chains of the human IgG molecule obtained by IEDB software. Most of the B-sheets in IgG molecules were located in 150 - 175 amino acid sequences of light chains and in 130-140, 160-170, 190-200, 220-245, 375-410 and 420-425 amino acid sequences of heavy chains respectively as was determined by IEDB software. Three epitopes sited to constant domain of IgG light chain (CL) were predicted. These epitopes were located at 150-175 and 180-205 amino acid sequences of light chain. Also three epitopes situated to second and third constant domains of IgG heavy chain (CH2 and CH3) were predicted. These epitopes were located at 200-270, 290-360 and 380-400 amino acid sequences of heavy chain. Based on the results of present study, 66% of human IgG light chain epitopes and 66% of human IgG heavy chain epitopes are situated at the IgG B-sheets locations. Also in present study, 91% of the amino acids forming B-sheets sited in the heavy chains of IgG molecule were located in the CH2 and CH3 domains and 100% of the human IgG heavy chain epitopes were located in the CH2 and CH3 domains Conclusion: According to the results of this study, most of human IgG epitopes are located in regions of the molecule where B-sheets are most likely situated, indicating a relatively high compatibility of B-sheets with IgG epitopes. Given that the B-sheets present in the second structure of proteins play an important role in epitope formation (6, 7), the occurrence of most human IgG epitopes (66%) at the location of B-sheets seems evident. Moreover, according to the findings of the present study, a total of 91% of the amino acids forming B-sheets sited in the heavy chains of IgG molecule were located in the CH2 and CH3 domains and 100% of the human IgG heavy chain epitopes were located in the CH2 and CH3 domains. This again indicates the high compatibility of B-sheets with epitopes located on the heavy chain of the human IgG molecule. The findings of our study are confirmed by Hajighasemi et al. study (31). In study of Hajighasemi et al. (31), about 91% of the linear epitopes located on the heavy chains of the IgG molecule, were situated in the CH2 and CH3 domains. Similarly, in our study 100% of human IgG heavy chain epitopes located on CH2 and CH3 domains. Also the results of present study are confirmed by another research (36) in which all (100% ) of spatial epitopes on fragment of crystallizable [Fc] component (part of heavy chains) of the human IgG molecule were located in CH3 and CH2 domains as were identified by DiscoTope and ElliPro immunoinformatic softwares. Similarly in the present study, 100% of IgG heavy chain epitopes were located in the CH2 and CH3 domains. Moreover the results of another study performed by Rohani et al. (37) support the findings of current study. Rohani et al., reported that most of the areas with highest surface accessibility, hydrophilicity and flexibility, were situated in CH2 and CH3 domains of human IgG heavy chains. (37). According to our results, most of B-sheets are located in the CH2 and CH3 domains of human IgG heavy chains. As B-sheets are regions with high accessibility, hydrophilicity and flexibility (6), presence of most B-sheets in the CH2 and CH3 domains which are reported to be highly accessible, hydrophilic and flexible ( 37), is reasonable. Furthermore, the high immunogenicity of B-sheets (39), once again confirms results of the present study. As immunogenicity plays an important role in epitope development, the high adaptation between the location of B-sheets and epitopes ( shown in our study) which both are highly immunogenic, seems logical. Besides since B-sheets play an important role in epitope formation(6, 7), the high compatibility between location of B-sheets and IgG epitopes ( shown in present study) seems sensible. Overall, presence the most of B-sheets and IgG epitopes in CL, CH2 and CH3 domains of human IgG, shows the high compatibility between location of B-sheets and epitopes in human IgG. These findings are precious tools not only for prediction of IgG immunogenic epitopes in order to producing specific monoclonal anti IgG antibodies to optimize current human IgG diagnostic kits but also for more exact detection of IgG functions. Moreover our results are helpful in producing of similar proteins for diagnostic and therapeutic aims, structure- function relationships and phylogenic studies. [ABSTRACT FROM AUTHOR]

Background and Objectives: Acute lymphocytic leukemia (ALL) is a type of cancer that affects white blood cells. The disease progresses rapidly mandating immediate treatment. ALL is the most common type of leukemia affecting children. An increase in the miR-222-3p molecule has been observed in the course of this disease. The microarray method can be used to examine the binding of this miRNA and target genes, which is not without limitations. Hence, the present study deployed computer and bioinformatics methods with different specialized software. Materials and Methods: Target genes were identified using different algorithms available in Mirbase, TargetScan, NCBI and RNAhybrid databases, and based on the scores obtained, the targets with the highest affinity were identified. Results: The findings of this study showed that the CDH4, AJAP1, HNRNPA0, UBN1 and ZNF618 genes are potential targets with higher specificity for miR-222-3p. Conclusions: Factors involved in selecting targets with high specificity and affinity is dependent on the amount of negative free energy and the number of base pairs formed between miRNA and 3'-UTR of different genes. [ABSTRACT FROM AUTHOR]

Introduction: COVID-19 is an acute respiratory infectious disease caused by the SARS-CoV-2 virus. There is an urgent need to discover antiviral drugs for better performance against new strains of coronaviruses (CoVs) due to the rapid spread of the disease despite scientific advances in vaccine development. This study aimed to evaluate the efficacy of quercetin and its analogues on the COVID-19 Mpro enzyme. Material & Methods: In this descriptive-analytical study, the threedimensional structures of quercetin analogues (20 compounds), standard drugs (ritonavir and lopinavir), and the COVID-19 Mpro enzyme were obtained from PubChem and PDB databases for bioinformatics study, respectively. Molecular docking studies of the compounds on the Mpro were performed using MOE-2014 software. Afterward, the physicochemical properties and biological activity of the compounds were predicted using Swiss ADME, PASS, and Swiss Target Prediction software. (Ethic code: IR.SKUMS.REC.1399.117) Findings: The findings of the present study showed that the most important bonds involved in drug-receptor binding are hydrogen, hydrophobic, and π-π interaction bonds. The best docking results were obtained for Baicalein, Genistein, Naringenin, and Quercetin compounds with strong binding energy (-12.83 to -13.54 kcal/mol), compared to ritonavir and lopinavir. These compounds have a greater tendency to bind to the catalytic amino acids His41 and Cys145 and other key amino acids of the active site of the COVID-19 Mpro enzyme. Discussion & Conclusion: Based on the results of bioinformatics studies, quercetin analogues had more effective inhibition than standard chemical drugs due to their suitable placement in the active site of the main protease enzyme of COVID-19 and can be good candidates for in vitro and in vivo studies. [ABSTRACT FROM AUTHOR]

Coronavirus disease 2019 (COVID-19) is an acute respiratory infection. Its virus called SARS-COV-2 which is an RNA virus with high homology to the bat coronavirus. In this review study, first the molecular and cellular characteristics and the proliferation and replication of SARS-COV-2 are investigated. Then, by reviewing bioinformatics studies regarding protected domain analysis, homology modeling, and molecular docking, the biological role of some specific SARS-COV-2 proteins are examined. The results showed that the open reading frame 8 (ORF8) and surface glycoprotein could bind to porphyrin. At the same time, ORF1ab, ORF10, and ORF3a can attack the heme part of hemoglobin to dissociate iron and form porphyrin. This attack reduces hemoglobin ability to carry oxygen and carbon dioxide. As a result, lung cells become severely inflamed due to their inability to exchange carbon dioxide and oxygen, which leads to large ground-glass opacities on CT scan images. Based on the bioinformatics results, chloroquine can prevent ORF1ab, ORF3a, and ORF10 from attacking hemoglobin to form porphyrin and avoid the binding of ORF8 and surface glycoprotein to porphyrin, which effectively relieves the symptoms of acute respiratory syndrome. In the current pandemic, bioinformatics studies are of great importance for preventing the spread of COVID-19, developing drugs and vaccines, and clinical practice. [ABSTRACT FROM AUTHOR]

Objective Acinetobacter baumannii is an important bacterium in causing nosocomial infections that has multidrug resistance. Several factors are involved in the resistance of these bacteria to drugs, the most important of which are the proteins OXA-24 beta-lactamase, OXA-23 beta-lactamase and OUTER MEMBRANE PROTEIN A (OMPA). In this regard, the aim of this study was to investigate the inhibitory effect of plant compounds in Salsify, Hyssop and Mangrove on these Acinetobacterproteins in silico. Materials and methods In this study, first the structure of plant compounds and proteins were obtained from PubChem and PDB databases, respectively. Then the physicochemical properties and mutagenic potential of plant compounds were predicted by Swiss ADME server and Toxtree-v2.6.13 software, respectively. ViewerLite, Chimera 1.14, Discovery Studio, AutoDockTools-1.5.6 and AutoDock Vina environments were used for molecular docking and interaction between plant compounds and proteins. The results were analyzed using three programs including AutoDockTools, Visualizer DS and Ligplot. Results According to mutagenicity potential and inhibition cytochromes results, compounds related to H. officinalis had higher mutagenic potential and cytotoxicity than the other two plants. In addition, the results showed that A. marina compounds have higher digestive absorption compared to other compounds. A comparative study of the interactions showed that the compounds of H. officinalis and A. marina have stronger interactions with the studied proteins. In addition, it was found that the strongest interactions occurred between plant compounds and 1BXW protein. On the other hand, T. graminifolius compounds were found to have weaker interactions with the evaluated proteins. Conclusions Based on docking results, among all compounds evaluated in all three proteins 1BXW, 3G4P and 4 K0X, the best docking results were related to cis-pinocamphone and 3,5-difluorophenyl ester 2,6-difluorobenzoic acid, respectively, from Hyssop and mangrove. In fact, these compounds with the most negative binding energy levels (-9 Kcal / mol and -8.8, respectively) have a greater tendency to bind to the key amino acids of the active site of all three proteins. Therefore, these compounds can be considered as important candidates for laboratory and in vivo testing of A. baumannii activity.

Objective DREB gene family is one of the effective transcription factors in plant tolerance to environmental stresses, including salinity stress. The aim of this study is to identify and evaluate the expression of DREB2 gene in different salinity concentrations in local wheat of Kalak Afghan. Materials and methods Salinity stress treatment was applied to wheat using NaCl salt. Experimental treatments included different salinity levels (0, 100, 150, 200, 250 and 300 mM). Then RNA extraction and cDNA synthesis were performed and the specific fragment of DREB2 gene was amplified with specific primers. The obtained transcript was then sent for sequencing and the sequence was analyzed to identify the DREB2 gene. Results Partial sequence of DREB2 gene of local wheat of Kalak Afghani was registered in NCBI database with access number KR106189. The results of sequencing showed that the obtained sequence is more than 95% similar to the sequence of this gene in Triticum dicoccoides and Agropyrum elongatum. Examination of the second structure of DREB2 protein sequence through PSIpred and SOPMA programs showed that this sequence has 46 alpha helices (32.86%), 11 beta helices (7.86%), 9 long strands (6.43%) and 74 is a random helix (52.86%). The results of evaluation of DREB2 gene expression showed that the concentration of 100 mM salinity treatment, has a significant effect on increasing gene expression. In general, the highest effect on DREB2 gene expression was 100 mM salinity treatment and the lowest value was 300 mM. Conclusions In general, the DREB2 gene increases plant tolerance compared to other stress-induced genes, which makes them a desirable target for genetic engineering and crop performance. It seems that the reason that the expression of DREB2 gene does not increase at high concentrations is that the salt load is greater than the cells' ability to transmit it. Under these conditions, salt is transferred to the cytoplasm and inhibits enzymatic activities. Also, a common consequence of the accumulation of high concentrations of salt is the accumulation of large amounts of Reactive oxygen species (ROS), which can cause protein oxidation, damage to cellular DNA, lipid peroxidation, and interaction with other vital cell components. Leads to cell toxicity.

Background: Hypertrophic cardiomyopathy is a common genetic cardiovascular disease with autosomal dominant inheritance and MYBPC3 gene has been frequently linked to its pathogenesis. Since, carriers of the 25 nucleotides deletion located on intron 32 of the MYBPC3 are at increased risk of heart disease we aimed to investigate this variant in hypertrophic cardiomyopathy patients and healthy population. Methods: DNA was extracted from 350 Blood samples including 42 hypertrophic cardiomyopathies and 308 healthy subjects and the region containing the deletion was amplified by PCR method. PCR products were analyzed on agarose gel and genotyping results were recorded. Results: Genetic counseling results revealed that 26.2% of patients were sporadic cases vs 59.5% with positive family history and there was a history of sudden cardiac death in the first degree relatives of 42.3% of the patients. Genotyping results showed that all samples had a single band of 198 bp, indicating no MYBPC3Δ25bp variant in HCM patients as well as 308 controls. Bioinformatics assessments revealed that MYBPC3Δ25bp had a frequency of 0.00438 on Iranome database with the highest incidence reported in the Baloch population. Conclusion: Since hypertrophic cardiomyopathy is related to sudden cardiac death, population studies in terms of predisposing factors are of particular importance. Our study results showed that MYBPC3Δ25bpshould not be considered as risk factor in the patients of northwest of Iran. However, according to the bioinformatics findings and reports of neighboring countries, it is suggested that MYBPC3Δ25bp to be studied in the eastern and southern Iranian hypertrophic cardiomyopathy patients.

Introduction: The prediction of microRNAs related to target genes using bioinformatics tools saves time and costs of the experimental analyses. In the present study, the prediction of microRNA target genes relevant to Alzheimer’s Diseases (AD) were compared with the experimentally reported data using different bioinformatics tools. Method: A total of 41 microRNAs associated with 21 essential genes involved in AD were selected based on experimental results reported in previously published literature. Then, the prediction of the target gene for each microRNA was done using three bioinformatics tools, including MirTarget, TargentScan, and Diana-microT. The results of the predictions for all three tools considering the reported target genes were compared with each other. Results: The results showed that MirTarget, TargetScan, and Diana-microT correctly predicted 66%, 61%, and 27% of microRNAs’ attachment to the previously reported target genes involved in AD, respectively. However, none of the tools could predict the attachment of 22% of the microRNAs to the target genes reported in the literature. Conclusion: It can be concluded that MirTarget and TargetScan can better predict the target gene for microRNAs involved in AD compared with Diana-microT. Considering the algorithm used in MirTarget, this bioinformatics tool provides more functional and accurate results in predicting the target genes for microRNAs and it is recommended for predicting the target genes of microRNAs. It can also be concluded that the reported target genes for microRNAs involved in AD need further investigations in some cases.

Introduction: Abortion is a common complication that refers to the early termination of pregnancy with the death of the fetus before the 20th week of pregnancy. Previous studies show that many genes are involved in this disease, including the CX3CR1 gene, which is one of the inflammatory response genes in the immune system. The pathogenicity of these variants was determined in this study using bioinformatics analysis. Material & Methods: In this study, the effects of rs3732378 and rs3732379 mutation were predicted using bioinformatics tools including SIFT, PolyPhen2, PROVEAN, Predict SNP, and Exome Variant Server. Changes in the stability of mutant proteins were investigated using IMutant and DynaMut tools. Moreover, modeling of the protein structure, docking, and protein-ligand interaction were performed using SWISSMODEL, SwissDock, and FRODOCK tools as well as PyMOL, Hawkdock, and MolSoft software, respectively. Findings: Many polymorphisms related to the CX3CR1 gene have been known to date. Out of 244 missense mutations in the dbSNP database, two variants (rs3732378 and rs3732379) have been reported in association with recurrent pregnancy loss related to the CX3CR1 gene. The results of bioinformatics analyses showed that both variants were predicted as pathogenic mutations and changed the stability of the protein structure and played a key role in interaction with the ligand. Discussion & Conclusion: The findings of this study indicate that two missense mutations in the CX3CR1 gene are an important candidate for recurrent miscarriage and their identification in patients with recurrent miscarriage can be regarded as a risk factor. [ABSTRACT FROM AUTHOR]

Introduction: A growing amount of evidence has revealed that circular RNAs (circRNAs), covalently closed single-stranded non-coding RNAs (ncRNAs), play significant roles in initiation and progression of breast cancer (BC) through different mechanisms. This study aims to investigate the hsa_circ_0003098 expression and its potential roles in BC, as well as the circRNA/miRNA/mRNA crosstalk in these contexts. Materials and Methods: Real-time PCR (RT-PCR) was used to assess the expression of hsa_circ_0003098 in the breast tumors compared with adjacent normal tissues. Its association with different demographic and clinicopathological characteristics of BC patients was assessed by statistical analysis. Finally, we used bioinformatic approaches to uncover the potential circRNA/miRNA/mRNA axes using some databases and software. Results: Our results revealed that hsa_circ_0003098 expression is significantly upregulated in BC tumors compared with the adjacent normal tissues. Moreover, hsa_circ_0003098 over-expression is significantly associated with lymph node metastasis and hair dye use. Next, we found that this circRNA can regulate key mRNAs and miRNAs in the ceRNA network. Conclusion: It is suggested that hsa_circ_0006960, which is significantly up-expressed in breast tumors and associated with lymph node metastasis, could play key roles in the physiopathology of breast cancer through sponging off the miRNAs involved in hsa_circ_0003098/miRNAs/mRNAs network. [ABSTRACT FROM AUTHOR]

Background and purpose: The aim of this study was to bioinformatically and experimentally evaluate the effect of melatonin on the expression levels of UBE2W and SSX2IP genes in melatonin treated Human Hepatocellular carcinoma G2 (HepG2) cancer cell line. UBE2W encodes a protein that promotes ubiquitination of Fanconi anemia complementation group proteins and may be important in the repair of DNA damage. SSX2IP belongs to an adhesion system, involved in cell movement and acts as a centrosome maturation factor. Materials and methods: At first, as a bioinformatics tool, MATLAB 2018a software was employed to evaluate the UBE2W and SSX2IP genes expression levels using microarray data. Then, after designing and preparing the primers, MTT and Real-time PCR were carried out in three replicates. Results: MTT assay showed that the viability of cancer cell significantly decreased in melatonin treated group (P≤0.01). The IC50 value was estimated to be 2080 µM for 48 hours, but no significant changes were seen in the survival of human normal cells (HUVEC-C) (P≥0.05). Real-time PCR proved that the expression levels of both genes increased (P≥0.05 in 24h and P≤ 0.05 in 48h) in melatonin treated cells compared to un-treated group, which was in accordance with bioinformatics analysis. Conclusion: Our study showed that UBE2W and SSX2IP genes could be used as therapeutic target genes for Hepatocellular carcinoma. However, complementary studies are needed to prove current findings.

Introduction: Long non-coding RNAs (lncRNAs) and miRNAs belong to a class of non-coding RNAs (ncRNAs) that play important roles and functions in the regulation of the expression of genes in main biological processes, such as cell proliferation, apoptosis, and differentiation. LncRNAs can potentially affect miRNAs in the forms of cis/trans to modulate their regulatory role. In this study, mRNA, miRNA, and lncRNA gene networks were predicted by web-based programs for three ectodermal pathway markers (BMP4, NOG, FGF8) in the mouse embryonic stem cells. Material & Methods: In this theoretical bioinformatics study, the miRNAs of the target genes (BMP4, NOG, and FGF8) were extracted and examined by MirWalk and TARGETSCAN databases to finally obtain the common miRNAs of these three genes. Following that, the target lncRNAs for common miRNAs were then extracted from the DIANA-Tool database. (Ethic code: 100/21560/2/پ( Findings: MiRs mmu-miR-92a-2-5p, mmu-miR-129b-5p, mmu-miR130b-5p, mmu-miR-692, mmu-miR-7009-3P, mmu-miR-7116-3p, and mmu-miR-7689-3p may affect the function of lncRNAs, including Kcnq1ot1, Gm26812, Gm4117, Gm11837, 4930423MO2Rik, Malat1, Gm12594, Gm3414, 5830444B04Rik, Gm2464, and NEAT1. Discussion & Conclusion: Due to the mutual relationships among lncRNA, miRNA, and mRNA, our results provided a novel perspective on lncRNAs for future research and experimental studies on ectodermal differentiation pathways and molecular mechanisms. [ABSTRACT FROM AUTHOR]

Background and Objectives Stem cell factor (SCF) is a blood cytokine that plays an important role in the differentiation of hematopoietic progenitor cells, as well as survival, proliferation, and differentiation of mast cells. Furthermore, it can enhance tumor cell proliferation and invasion. Recombinant or natural form of leucine zipper can cause homo- or heterodimerization of many types of proteins which are attached to it and have a favorable effect on their function or stability. The present study investigates the expression of leucine zipper-bound SCF. Materials and Methods This experimental study consists of designing a recombinant protein by bioinformatic softwares and the laboratory part of the study. The indices of interest for designing of the protein structures were analysed by various softwares including PSIpred and Chimera and the recombinant protein stability by RAMPAGE. The sequence synthesis was performed after determination of the enzymatic cleavage sites with NEBcutter. Then, codon optimization for Origami was done with Optimizer. Cloning in TOP10 and the protein expression in Origami were performed. Results The results of bioinformatics studies on the Leucine Zipper/SCF recombinant protein included the necessary criteria for the possible native structure of the protein and the high score for viability in the living cell. Furthermore, the results of the experimental section showed successful cloning and expression. Conclusions In the present study, the results indicated the successful cloning and expression of the recombinant Leucine Zipper/SCF protein. [ABSTRACT FROM AUTHOR]

In this paper, we consider a three parametric regularly varying generalized Hypergeometric distribution which have been generated by Birth-Death process for describing phenomena in bioinformatics (Danielian and Astola, 2006). Under satisfying some conditions, we obtain the system of likelihood equations which its solution coincides with the maximum likelihood estimators. The given maximum likelihood estimators are the same as some moment estimators. Moreover, an approximate computation of the maximum likelihood estimations for the unknown parameters is given. Using MCMC, simulation studies are proposed. Finally, in order to present applications, some real data sets in bioinformatics are fitted with the model. Based on some important criterions, this model is compared with four other discrete distributions in bioinformatics. We see that the generalized Hypergeometric distribution provides a better fit than four other discrete distributions../files/site1/files/61/9(1).pdf

Objective Cadmium is a highly toxic and widespread soil pollutant threatening human and animal. Plants effectively help to eliminate environmental pollution by up taking of heavy metals and tolerate cadmium stress through a variety of mechanisms, but biochemical pathways and genes involved in the response of plants to cadmium stress have not been fully and comprehensively identified. Materials and methods Following proteomic studies on the Arabidopsis thaliana mutant in which AT2G37050 (receptor like kinase gene) was knocked-out, it was identified that 150 proteins that were present in the wild (control) plant have been disappeared in the mutant plant. In current study, biological function of AT2G37050 gene/GO terms has been investigated by GeneMANIA and agriGO algorithms. GO term is a controlled vocabulary system describing biological entities in three aspects (biological process, molecular function, and cellular component) in different organisms. Results Bioinformatics studies resulted from the GeneMANIA algorithm showed that the AT2G37050 gene is involved in the biological process of response to cadmium ion. The agriGO algorithm was then used to study GO terms at three levels of biological process, molecular function and cellular component, and role of the AT2G37050 gene and biological process of response to cadmium ion was reconfirmed. In addition, significant GO terms (FDR

مقدمه: عمق و گستردگی اطلاعات موجود در جامعه علمی و تحقیقاتی کنونی، فرصت خوبی را برای بهره‌برداری از این اطلاعات فراهم آورده است. در این میان، حوزه بیوانفورماتیک به عنوان یک زمینه علمی به نسبت جدیدی که نقش مهمی در مطالعات زیستی دارد، توجه زیادی از رشته‌های مختلف مانند فن‌آوری اطلاعات، ریاضیات و علوم زیستی نوین را در سال‌های اخیر به خود جلب کرده است. در این راستا، هدف از انجام پژوهش حاضر، تعیین وضعیت مقالات منتشر شده در زمینه‌ بیوانفورماتیک بر اساس پایگاه Clarivate بود. روش بررسی: این مطالعه توصیفی به روش علم‌سنجی انجام ‌شد. جامعه آماری شامل تمامی مقالات حوزه بیوانفورماتیک در پایگاه Clarivate طی سال‌های 1970 تا 2018 بود. داده‌ها با استفاده از فراوانی و درصد توصیف گردید. یافته‌ها: روند رشد مقالات بیوانفورماتیک در پایگاه Clarivate از سال 1970 تا 2018 سیر صعودی داشت. مجموع مقالات مربوط به این سال‌ها، 806/53 عنوان بود. در این مقالات، حوزه‌های موضوعی بیوشیمی مولکولی زیستی، کامپیوتر، بیوتکنولوژی، ژنتیک و زیست‌شناسی محاسباتی به عنوان فعال‌ترین حوزه‌ها شناسایی گردید. ایالات ‌متحده آمریکا، چین، آلمان، انگلستان و کانادا از جمله فعال‌ترین کشورهای تولیدکننده مقالات محسوب شدند. بیشترین مقالات به Zhang Y، Wang Y، Wang J، Li Y و Li J اختصاص داشت. دانشگاه‌های کالیفرنیا، هاروارد و آکادمی علوم چین بیشترین مقالات این حوزه را منتشر کرده بودند. بالاترین تعداد مقاله در مجله PLOS ONE چاپ شده بود. نتیجه‌گیری: از بین 164 کشور، ایران در رتبه 22 قرار داشت که این وضعیت نسبت به جایگاه جهانی مطلوب است و با توجه بیشتر به انجام مطالعات در این زمینه، می‌توان به وضعیت بهتری رسید.

Introduction1 Obviously, the recent decades strategy in cancer therapy, anticancer drug discovery and drug improvement is to characterize, distinguish and validate the most promising cancer-related molecular targets to which new drugs can be designed. The unique features of the pancreatic RNase (RNase A) such as high activity, stability, lack of a cofactor, and small molecular size have made it the most popular enzyme in the ribonuclease family. Specifically, RNase A is involved in endonucleolytic cleavage of 3'-phosphomononucleotides and 3'- phosphooligonucleotides ending in C-P or U-P with 2', 3'- cyclic phosphate intermediates. RNase A was purified from the Bovidae family including bovine, ovine, bison, eland, goat and gnu. Although phylogenetic analyses of RNase A revealed high similarity among members of the Bovidae family, some functional mutations were also found. Several studies showed that the RNA hydrolyzing action of ribonuclease is able to induce apoptosis and cell death in cancer cells, independently. This effect could be enhanced thousands of times when ribonuclease is linked to antibodies. These enzymes show potent cytotoxic activity on cell internalization but do not show sensible immunogenicity or non-specific toxicity toward normal cells. Ovine pancreatic ribonuclease enzyme is a member of super family RNase A, it can be a good candidate as a toxin for designing new drugs. The objective of this study was to produce ovine pancreatic ribonuclease enzyme in E. coli and characterize its activity. Materials and methods All structures needed for this study were downloaded from the Protein Data Bank (PDB) website (http://www.rcsb.org/). PDB files (accession numbers: 10.2210/pdb1YV6/pdb and 10.2210/pdb3SNF/pdb) related to natural Bovine Pancreatic RNase were selected. Gene synthesis and production of recombinant protein were conducted by using the pelB signal sequence at the beginning of the structure for periplasmic protein production. The native ovine RNase and bovine RNase A were optimized for E. coli host by GenRay codon optimization service and sent to GenRay Biotechnology (Shanghai, China) for synthesis. The target genes in pGH vector were sub-cloned in pUC19 and then cloned into the pET21b (+) vector between XbaI and HindIII sites. After transformation, E. coli cells containing recombinant pET21b (+) were cultivated in LB broth medium containing ampicillin. To extract the proteins, osmotic shock methods were applied. After that Q-sepharose chromatography was used to extract the target protein. Finally, Bradford analysis was used to determine the protein concentration. The ribonucleolytic activity of the recombinant native ovine RNase A was compared with native bovine RNase A following Tripathy et al (2013) method. To investigate the antitumor activity of recombinant proteins, HeLa cells were prepared for seeding in a 96-well flask and incubated at 37°C in 5% CO2 for 72 h. Results and discussion The production of native ovine and bovine RNase A was confirmed by SDS PAGE. Protein purification was successfully performed using osmotic shock in the Q-Sepharose column. Although our findings confirmed protein expression, no detectable proteins other than RNase A was observed in the LB medium, indicating that almost all the proteins were expressed either inside the bacterial cell or secreted into the periplasmic area. According to the Bradford analysis, the concentration of the recombinant proteins extracted was 4.78 ovine RNase A. Based on our results, it was shown that the ribonucleolytic activity of native ovine RNase A was 558±22. The results showed that RNase A exhibited resistance to pepsin degradation during the whole incubation process (22 h), in the course of which 90% of RNase A remained undamaged. To determine the cytotoxic effect of ovine RNase on the HeLa cell line, MTT assay was done following incubation with ovine RNase A. The commercial RNase A was used as control. The results showed that ovine RNase A and bovine RNase A had no cytotoxic effect on HeLa cells. When RNase treatment was done by the lipofectamine 3000 (Thermo, USA), the cytotoxicity effect was observed. Several studies have shown that some ribonucleases such as onconase, bovine seminal ribonuclease and bovine pancreatic ribonuclease have a great promise as cancer immunotherapeutic agents and cause a significant reduction in the protein synthesis of tumor cells after internalization into cytosol. Conclusion Our findings demonstrate that ovine RNase similar to bovine RNase has a great potential for use in drug design industry. We revealed that the native ovine RNase A was more stable than the native bovine RNase. In future work, we intend to fuse the engineered ovine-RNase A to dedicated recombinant antibodies for cancer therapy and investigation of engineered immuno-ribonuclease potency and cell killing effects as a fusion protein. [ABSTRACT FROM AUTHOR]

Allium includes more than 920 species worldwide. Iran is the main habitat of many wild Allium species. In this study, onion tissue of two species A. stipitatum and A. scabriscapum from two subspecies Melanocrommyum and Reticulatobulbosa were used to evaluate and identify the expression pattern of storage proteins obtained by the SDS-PAGE method. Twodimensional gel electrophoresis and MALDI TOF/TOF MS mass spectrometry were used to identify protein spots and to observe the expression changes of the two species. Analysis of gels by two-dimensional electrophoresis revealed 138 protein spots. The results showed a slight similarity between the gels and a different expression pattern between the samples, based on which a small number of similar spots were obtained in both gels. The results showed that there were significant differences in the expression profile of onion tissue proteins of these samples. Using the T-test, 9 protein spots with the most significant changes were identified at the 5% probability level, and among them, 3 different spots were identified using Mass spectrometry was selected for identification. The results of mass spectrometry experiments and matching of the obtained data with NCBI and Uniprot databases and bioinformatics tools showed that the tested spots were consistent with Maturasek proteins, Agmatine coumaroyltransferase-1, and a protein with the unknown function called Hypothetical Protein. The findings of this study showed that since the samples belonging to A.stipitatum and A.scabriscapum are of the same genus, but the expression pattern of their stored proteins in onion tissue is very different from each other. [ABSTRACT FROM AUTHOR]

Strawberry belongs to the genus Fragaria and family Rosaceae. Now days, the species Chiloensis, Virginia, and hybrid between them, namely, Ananasae, are cultivated. This perennial herb has a small genome that is prone to the transfer of Fragaria ananasae cultivated strawberry genes and other important economic plants from the Rosaceae family. Present study was conducted to study and compare the complete chloroplast sequence of Fragaria vesca, analyses of its genome structure, gene content and organization, repeated sequences, codon usage and comparison with two cultivated octaploid sequenced Fragaria species (F.chiloensis and F.virginiana). F.vesca chloroplast (cp) genome is 155691 bp in length with conserved quadripartite structure. Single copy region of cp genome is separated by the two inverted regions. The large single copy region is 85561 bp, and the small single copy region is 20204 bp whereas the inverted repeat is 24963 bp each. The plastidic genome has 130 single genes and 18 duplicated genes located in inverted repeats. The singletons encode 85 proteins, 8 ribosomal RNA genes and 37 transfer RNA genes. Amongst all plastidic genes only 18 genes appeared to have 1-2 introns and when compared with DNA of two cultivated octaploeids, rps18 was the only duplicated gene in F.vesca and rps 12, rpl 2 and trn I-GAU are genes which are located in inverted repeats. Despite the high level of conservation in chloroplast SSRs, these are useful in analysis of genetic diversity due to their greater efficiency as opposed to genomic SSRs.