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Antagonistic interactions among different functional guilds of nematodes have been recognized for quite some time, but the underlying explanatory mechanisms are unclear. We investigated responses of tomato (Solanum lycopersicum) to two functional guilds of nematodes - plant parasite (Meloidogyne javanica) and entomopathogens (Heterorhabditis bacteriophora, Steinernema feltiae belowground, and S. carpocapsae) - as well as a leaf mining insect (Tuta absoluta) aboveground. Our results indicate that entomopathogenic nematodes (EPNs): 1) reduced root knot nematode (RKN) infestation belowground, 2) reduced herbivore (T. absoluta) host preference and performance aboveground, and 3) induced overlapping plant defense responses by rapidly activating polyphenol oxidase (PPO) and guaiacol peroxidase (GP) activity in roots, but simultaneously suppressing this activity in aboveground tissues. Concurrently, we investigated potential plant signaling mechanisms underlying these interactions using transcriptome analyses. We found that both entomopathogens and plant parasites triggered immune responses in plant roots with shared gene expression. Secondary metabolite transcripts induced in response to the two nematode functional guilds were generally overlapping and showed an analogous profile of regulation. Likewise, we show that EPNs modulate plant defense against RKN invasion, in part, by suppressing active expression of antioxidant enzymes. Inoculations of roots with EPN triggered an immune response in tomato via up-regulated phenylpropanoid metabolism and synthesis of protease inhibitors (PIs) in plant tissues, which may explain decreased egg laying and developmental performance exhibited by herbivores on EPN-inoculated plants. Furthermore, changes induced in the volatile organic compound (VOC)-related transcriptome indicated that M. javanica and/or S. carpocapsae inoculation of plants triggered both direct and indirect defenses. Our results support the hypothesis that plants 'mistake' subterranean EPNs for parasites, and these otherwise beneficial worms activate a battery of plant defenses associated with systemic acquired resistance (SAR) and/or induced systemic resistance (ISR) with concomitant antagonistic effects on temporally co-occurring subterranean plant pathogenic nematodes and terrestrial herbivores.

One of the most promising ways to diagnose cancer especially colorectal cancer (CRC) is to trace its epigenetic events. In this article, a discovery step for detection of methylated DNA markers (MDMs) was performed using SureSelectXT Methyl-Seq in CRC case and control groups in addition to several methylation profiling datasets (GSE48684, GSE53051, GSE77718, GSE101764, and GSE42752). In silico validation of MDMs in colorectal and other cancers was conducted by Lnc2met. MethyLight assay was run on 40 and 47 case and control formalin-fixed paraffin-embedded tissues, respectively and the performance of selected genes were classified by support vector machine (SVM). As a result, 180 regions were identified among all common genes. In addition to SEPT9 and SFRP2, the best three MDM regions were selected from SLC30A10, AKR1B1 and GALNT14. Based on all assays, the best performance was accomplished by SEPT9/AKR1B1 with 98% sensitivity, 99% specificity, 125 positive likelihood ratio, 0.02 negative likelihood ratio and 5074 diagnostic odds ratio. Our results indicate that the AKR1B1/SEPT9 methylation panel detects CRC with a higher performance than SEPT9 methylation, which is a commercial diagnostic test for CRC. However, the creation of a clinically valuable test derived from this study requires performance evaluation in liquid biopsies.

DNA methylation is a fundamental epigenetic process and have a critical role in many biological processes. The study of DNA methylation at a large scale of genomic levels is widely conducted by several techniques that are next-generation sequencing (NGS)-based methods. Methylome data revealed by DNA methylation next-generation sequencing (mNGS), should be always verified by another technique which they usually have a high cost. In this study, we offered a low-cost approach to corroborate the mNGS data. In this regard, mNGS was performed on 6 colorectal cancer (case group) and 6 healthy individual colon tissue (control group) samples. An R-script detected differentially methylated regions (DMRs), was further validated by high resolution melting (MS-HRM) analysis. After analyzing the data, the algorithm found 194 DMRs. Two locations with the highest level of methylation difference were verified by MS-HRM, which their results were in accordance with the mNGS. Therefore, in the present study, we suggested MS-HRM as a simple, accurate and low-cost method, useful for confirming methylation sequencing results.

The insulin-like growth factor (IGF) axis is fundamental for mammalian growth and development. However, no comprehensive reference data on gene expression across tissues and pre- and postnatal developmental stages are available for any given species. Here we provide systematic promoter- and splice variant specific information on expression of IGF system components in embryonic (Day 48), fetal (Day 153), term (Day 277, placenta) and juvenile (Day 365-396) tissues of domestic cow, a major agricultural species and biomedical model. Analysis of spatiotemporal changes in expression of IGF1, IGF2, IGF1R, IGF2R, IGFBP1-8 and IR genes, as well as lncRNAs H19 and AIRN, by qPCR, indicated an overall increase in expression from embryo to fetal stage, and decrease in expression from fetal to juvenile stage. The stronger decrease in expression of lncRNAs (average -16-fold) and ligands (average -12.1-fold) compared to receptors (average -5.7-fold) and binding proteins (average -4.3-fold) is consistent with known functions of IGF peptides and supports important roles of lncRNAs in prenatal development. Pronounced overall reduction in postnatal expression of IGF system components in lung (-12.9-fold) and kidney (-13.2-fold) are signatures of major changes in organ function while more similar hepatic expression levels (-2.2-fold) are evidence of the endocrine rather than autocrine/paracrine role of IGFs in postnatal growth regulation. Despite its rapid growth, placenta displayed a more stable expression pattern than other organs during prenatal development. Quantitative analyses of contributions of promoters P0-P4 to global IGF2 transcript in fetal tissues revealed that P4 accounted for the bulk of transcript in all tissues but skeletal muscle. Demonstration of IGF2 expression in fetal muscle and postnatal liver from a promoter orthologous to mouse and human promoter P0 provides further evidence for an evolutionary and developmental shift from placenta-specific P0-expression in rodents and suggests that some aspects of bovine IGF expression may be closer to human than mouse.

Genomic imprinting results in monoallelic expression of genes in mammals and flowering plants. Understanding the function of imprinted genes improves our knowledge of the regulatory processes in the genome. In this study, we have employed classification and clustering algorithms with attribute weighting to specify the unique attributes of both imprinted (monoallelic) and biallelic expressed genes. We have obtained characteristics of 22 known monoallelically expressed (imprinted) and 8 biallelic expressed genes that have been experimentally validated alongside 208 randomly selected genes in bovine (Bos taurus). Attribute weighting methods and various supervised and unsupervised algorithms in machine learning were applied. Unique characteristics were discovered and used to distinguish mono and biallelic expressed genes from each other in bovine. To obtain the accuracy of classification, 10-fold cross-validation with concerning each combination of attribute weighting (feature selection) and machine learning algorithms, was used. Our approach was able to accurately predict mono and biallelic genes using the genomics and proteomics attributes.

Aslaminejad, A.A., Nassiry, M.R., Farajollahi, H., Mahdavi, M., Abbasi, H. and Javadmanesh, A. 2010. Polymorphism in exon-3 of leptin gene in iranian native cattle breeds. J. Appl. Anim. Res., 37: 225–228. This study describes genetic variability of the leptin gene in four Iranian native cattle breeds. A 317 bp fragment of exon 3 of the leptin gene from 332 individuals in four different populations of Iranian cows including Golpayegani (N=92), Najdi (N=54), Sarabi (N=82) and Sistani (N=104) was amplified and PCR products digested with Nrul restriction enzyme. Allele C in Golpayegani, Najdi, Sarabi and Sistani breeds were 75, 78, 59 and 52 per cent, respectively. Observed heterozygosities were 0.29, 0.37, 0.68 and 0.92 for Golpayegani, Najdi, Sarabi and Sistani cattle, respectively. This study suggests that allele frequencies of leptin differed among Iranian cattle breeds.

This study describes genetic variability in the BoLA-DRB3 gene in Iranian native cattle (Bos Indicus and Taurus) and relationships between these breeds. This is the first study of genetic polymorphism of the BoLA-DRB3 gene in Iranian native cattle. We examined exon 2 of the major histocompatibility complex (MHC) class II DRB3 gene from 203 individuals in four populations of Iranian native cattle (52 Sarabi, 52 Najdi, 49 Sistani, 50 Golpayegani cattle) using the hemi-nested PCR-RFLP method. We identified the 36 previously reported alleles and one novel pattern (*eac). Analysis of the frequencies of the various BoLA-DRB3.2 alleles in each breed indicated that DRB3.2*52 in Sarabi cattle (23%), DRB3.2 *14 and *24 alleles in Najdi cattle (13%), DRB3.2 *8 allele in Sistani cattle (22%) and DRB3.2*16 allele in Golpayegani cattle (14%), were the most frequent alleles. Allelic frequencies ranged from 1 to 23% among the 36 alleles and there were some alleles that were found only in Iranian cattle. Effective number of alleles in the four breeds was estimated to be 7.86, 11.68, 7.08 and 3.37 in Sarabi, Najdi, Sistani and Golpayegani, respectively. Observed heterozygosities were the highest in Sarabi (94%) and Najdi (94%). A population tree based on the frequency of BoLA-DRB3.2 alleles in each breed suggested that Najdi, Sarabi and Golpayegani cattle clustered together and Najdi and Sarabi were the closest breeds. Sistani cattle differed more from these three breeds. These new data suggest that allele frequencies differ between Iranian cattle breeds.

The genotypes for Beta-Lactoglobulin (BLG) and calpastatin (CAST) were determined by Polymerase Chain Reaction (PCR) and restriction enzyme digestion and genotyped for calpain (CAPN) by PCR-SSCP method in an Iranian breed sheep, Kurdi. Blood samples were collected from 100 pure bred Kurdi sheep from Kurdi breeding station located in Shirvan, Mashhad. The extraction of genomic DNA was based on Guanidin Thiocyanate-Silica gel method. After PCR reaction, amplicons were digested with restriction enzymes MspI and RsaI for beta-lactoglobulin and calpastatin genes, respectively. The beta-lactoglobulin locus had 3 genotypes with frequencies of 0.24, 0.54 and 0.22 for AA, AB and BB, respectively; calpastatin genotypes had 2 genotypes with frequencies of 0.76 and 0.24 for MM and MN genotypes, respectively. Calpain genotypes were analyzed with SSCP method, which had 2 genotypes with frequencies of 0.92 and 0.08 for AA and AB, respectively. Heterozygosity value for beta-lactoglobulin locus was 49% and for calpastatin and calpain loci was very low (24 and 8%, respectively). chi2 test confirmed the Hardy-Weinberg equilibrium for three loci in this population. These data provide evidence that Iranian's Kurdi sheep breed have a variability, which opens interesting prospects for future selection programs, especially marker-assistant selection between different genotypes of different locus and milk and cheese characteristics, gain and meat traits and also for preservation strategies.

BLAD is a hereditary disease in Holstein dairy cattle. The defective allele of CD18 gene which is responsible for this disease has a recessive inheritance. The recessive homozygous form (BL/BL) is lethal and since carrier animals have viability, BLAD frequency increases by use of carrier bulls in Artificial Insemination (AI). BLAD carriers can be detected easily by means of polymerase chain reaction followed by restriction analysis of the amplicons. In this study DNA samples from Holstein (n = 30) and Brown Swiss (n = 10) bulls from Abbas Abad AI center (Khorasan state of Iran) were analysed. A 101 bp fragment from the polymorphic region of CD18 gene located on chromosome 1 was amplified by PCR. Restriction enzymes TaqI and HaeIII were used to identify genotypes. Digestion products were screened by electrophoresis on 8% non-denaturing polyacrylamide gel and visualized by ethidium bromide staining. Frequencies of BL/TL (carrier) genotypes in Holstein and Brown Swiss bulls were 3.33% and 0%, respectively. Our pedigree studies of the carrier bull in this experiment revealed that the mutation inherited to him from Hawkeye bull (CANM 369995, BL). Although the elimination of BLAD-carrier bulls from the Holstein world would be the most efficient method to control this genetic disorder, many BLAD-carrier bulls are still listed commercially for AI and BLAD is still occurring in Iran. Monitoring the prevalence of BLAD-carriers in random selected herds may be helpful in judging the effectiveness of the BLAD-control program.

Identification of animal species used in commercial meat products is important with respect to economic and sanitary issues. The aim of this research was to realize ruminant species in meat products using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay. The universal CB7u primers pair was used for amplifying ~195 bp fragment from a variable region of cytochrome- b mitochondrial DNA gene by polymerase chain reaction. Species differentiation was realized by digestion of the amplified ~195 bp fragments with Sse9I restriction enzyme. The results indicate that 7/7 of Kebab loghmeh, 9/10 of minced meat, 4/8 of beef burger and 2/5 samples of canned stew samples, were contaminated with one of prohibited ruminant species residual. Furthermore, the results reveal that 5/30 of samples had cross-contamination with a mixture of meat originated from various species, which was against the labelled nutrition information. Our results indicate that the PCR-RFLP technique is a powerful and reproducible test for detection and separation of ruminant species residuals in commercial meat products, especially in developing countries. Keywords: Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), mitochondrial DNA, ruminant species, commercial meat products, cytochrome- b gene

Candidate gene approaches facilitate discovering and localizing causative genes for quantitative traits and polymorphisms within selected candidate genes can be tested for association with variationin the quantitative trait (Campbell et al., 2003). Leptin is a 16-kDa protein that is synthesized and secreted predominantly by white adipocytes tissue and its expression is regulated by body fatness and energy balance. Leptin gene expressed in a variety of tissues including adipose tissue, placenta, mammary glands, skeletal muscles, gastric mucosa, brain and pituitary glands. In cattle, the leptin gene is located on chromosome 4 and consists of three exons. Several studies shown the association between polymorphism at the leptin gene and feed intake, energy balance, fertility and immune functions. It has been shown that leptin gene influences milk performance in cattle and reproduction in beef cattle(Liefers et al., 2002). Buchanan et al. (2002) described a cytosine (C) to thymine (T) substitution in exon 2 of the leptin gene of the B. taurus breeds, suggesting the existence CC, TT and CT genotypes. The aim of this study was to identify the single nucleotide polymorphism of exon 2 in Iranian Taleshi cattle.

DNA-based molecular methods have made possible genotyping of animals of any age and sex for milk and meat genes, thus providing a potentially more efficient and flexible selection tool. Among specific genes that may affect economically important traits in sheep, the Beta-lactoglobulin (BLG) locus has been extensively studied [Barillet et al., 2005]. The genotype BB of BLG seems to be associated with higher milk yield; on the other hand genotypes AA and AB seem to be superior in protein and casein content and crude yield [Garzon et al., 1992]. Another genes intensively investigated in farm animal are that of calpastatin (CAST) and calpain (CAPN). Calpastatin and calpain (CAPN) deserves special attention because of their major role in meat production. The calpain-calpastatin system (CCS) comprises a family of calcium dependent neutral proteases. The CAPNs have been shown to play the major role in post mortem tenderization in beef, lamb, and pork by degrading specific muscle structural proteins. The aim of the present study was to identify genotypes of BLG, CAST and CAPN genes in Kurdi sheep breed by PCR.