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Background: Litter size in mice is an important fitness and economic feature that is controlled by several genes and influenced by non-genetic factors too. High positive selection pressure in each generation for Litter size at birth (LSB), resulted in the development of high and low prolific lines of inbred Swiss albino mice (SAM). Despite uniform management conditions, these lines showed variability in LSB across the generation., Methods and Results: Variation in estrous-phased ovarian gene expression between high (LSB ≥ 12) and low prolific lines (LSB ≤ 3) of F 4 inbred SAM, was explored using RNA-Seq. Estrous phase assessment was done using vaginal cytology. A total of 870 differentially expressed genes (DEGs) were identified; among which, 287 genes were significantly up-regulated while 583 genes were down-regulated in HLS as compared to the LLS group. DEGs were assigned to 166 Gene Ontology (GO) terms and KEGG pathways. In HLS, the significantly up-regulated DEGs were involved in ovarian cell-cell signaling, regulation of biological activity and ovarian metabolic-associated pathways. Most down-regulated DEGs were expressed in immune-related pathways, indicating that immunological dampening is associated with a high ovulation rate and higher level of progesterone concentration leading to physiological changes responsible for higher fecundity., Conclusions: The present study, based on bulk RNA-seq analysis reflects the aggregate gene expression of the whole ovarian tissue, and reveals 24 DEGs that could be used as candidates for litter size attributes in future polymorphism and functional studies to gain further insights into the mechanisms underlying litter size variations in animals., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2025. The Author(s), under exclusive licence to Springer Nature B.V.)

Cerebral ischemic stroke ranks among the leading causes of death and disability worldwide. A significant challenge, beyond the lack of effective therapies, is the frequent oversight of sex as a vital factor in stroke research. This study focuses on elucidating the sex-specific epigenetic mechanisms that contribute to neural damage and recovery in cerebral ischemia. In our previously reported study, we demonstrated that following ischemia-induced cerebral artery occlusion (ICAO), female striatal tissue exhibited an early reinstatement of H3K9me2 marks on the promoters of inflammatory genes compared to male striatal tissue. This restoration led to a reduction in the expression of inflammatory cytokines, ultimately contributing to accelerated recovery in females. Building upon these findings, the current study aimed to investigate the unidentified molecular pathways responsible for the accelerated recovery observed in females. To explore this, we performed illumina-RNA sequencing on striatal tissues 24-h post-ICAO. Interestingly, our analysis revealed differential regulation of H3K27me2 marks on the promoters of various neurogenic genes at an early stage, which facilitated early neurogenesis in the female striatum. This investigation identifies an epigenetic modulator, kdm6b/jmjd3, targeting H3K27, and delineates its sex-specific role in neural stem cell proliferation. The findings contribute to a comprehensive model linking gender-specific epigenetic regulation, neurogenesis, and post-ICAO recovery. In conclusion, the identified epigenetic modulators and their roles in neurogenesis offer potential targets for refined therapeutic interventions, emphasizing the importance of personalized and sex-specific considerations in stroke studies., Competing Interests: Declarations. Competing interests: The authors declare that they have no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Grass carp hemorrhagic disease is caused by the grass carp reovirus (GCRV). The disease spreads rapidly and has a high fatality rate, which seriously affects grass carp culture. Moreover, the molecular mechanisms underlying grass carp hemorrhagic disease remain unclear. To decipher the effects of GCRV on grass carp tissues, resistant grass carp A (GA) and susceptible grass carp B (GB) were selected through GCRV treatment, and control grass carp C (GC) was also established. The gill, liver, and muscle tissues exhibited different onset symptoms under the influence of GCRV by histological observation. We selected muscle samples with significant differences in symptoms for Illumina RNA sequencing. Analyses using Gene Ontology and Kyoto Encyclopedia of Genes and Genomes revealed 3512, 3074, and 1853 differentially expressed genes between "GC vs. GB," "GC vs. GA," and "GA vs. GB," respectively. Additionally, 40 differential immune-related genes and 28 differential interferon-stimulating genes (ISGs) related to the interferon (IFN) pathway were identified. The expression of immunogene-related genes of GB and GA, such as MDA5, IL-34, NF-KB, TRIM25, SOCS3, CEBPB, and BCL2, and genes associated with the JAK-STAT signaling pathway, such as IRF4, STAT1, STAT3, JAK 1, and JAK 2, was significantly upregulated. The IFN and JAK-STAT signaling pathways were closely related to anti-GCRV infection. The transcriptome data and predicted immune genes and ISGs in this study provide novel insights into the treatment of GCRV., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Opisina arenosella is one of the main pests harming coconut trees. To date, there have been few studies on the molecular genetics, biochemistry and physiology of O. arenosella at the transcriptional level, and there are no available reference genomes. Here, Illumina RNA sequencing combined with PacBio single-molecule real-time analysis was applied to study the transcriptome of this pest at different developmental stages, providing reference data for transcript expression analysis. Twelve samples of O. arenosella from different stages of development were sequenced using Illumina RNA sequencing, and the pooled RNA samples were sequenced with PacBio technology (Iso-Seq). A full-length transcriptome with 41,938 transcripts was captured, and the N50 and N90 lengths were 3543 bp and 1646 bp, respectively. A total of 36,925 transcripts were annotated in public databases, 6493 of which were long noncoding RNAs, while 2510 represented alternative splicing events. There were significant differences in the gene expression profiles at different developmental stages, with high levels of differential gene expression associated with growth, development, carbohydrate metabolism and immunity. This work provides resources and information for the study of the transcriptome and gene function of O. arenosella and provides a valuable foundation for understanding the changes in gene expression during development. • For the first time, PacBio Iso-Seq and RNA-seq were combined to analyze the Opisina arenosella transcriptomes. • Gene expression profiles were obtained by mapping RNA-seq short sequences to the full-length transcript. • Significant differences in gene expression profiles were found, especially growth and development. • The transcriptomes results help understand the changes in gene expression during the development of Opisina arenosella. [ABSTRACT FROM AUTHOR]

Bulblet initiation, including adventitious bud initiation and bulblet formation, is a crucial process for lily and other bulbous flowers that are commercially propagated by vegetative means. Here, by a hybrid strategy combining Pacific Biosciences (PacBio) full-length sequencing and Illumina RNA sequencing (RNA-seq), high-quality transcripts of L. brownii (Lb) and its variety, L. brownii var. giganteum (Lbg), during in vitro bulblet initiation were obtained. A total of 53,576 and 65,050 high-quality non-redundant full-length transcripts of Lbg and Lb were generated, respectively. Morphological observation showed that Lbg possessed a stronger capacity to generate bulblets in vitro than Lb, and 1 mg L−1 2,4-dichlorophenoxyacetic acid (2,4-D) significantly increased bulblet regeneration rate in two lilies. Screening of differentially expressed transcripts (DETs) between different stages and Mfuzz analysis showed 0 DAT to 1 DAT was the crucial stage with the most complex transcriptional change, with carbohydrate metabolism pathway was significantly enriched. In addition, 6,218 and 8,965 DETs were screened between the 2,4-D-treated group and the control group in Lbg and Lb, respectively. 2,4-D application had evident effects on the expression of genes involved in auxin signaling pathway, such as TIRs, ARFs, Aux/IAAs, GH3s and SAURs. Then, we compared the expression profiles of crucial genes of carbohydrate metabolism between different stages and different treatments. SUSs, SUTs, TPSs, AGPLs, GBSSs and SSs showed significant responses during bulblet initiation. The expression of CWINs, SUTs and SWEETs were significantly upregulated by 2,4-D in two lilies. In addition, 2,4- D increased the expression of starch degradation genes (AMYs and BAMs) and inhibited starch synthesis genes (AGPLs, GBSSs and SSs). SBEs were significantly upregulated in Lbg but not in Lb. Significant co-expression was showed between genes involved in carbohydrate metabolism and auxin signaling, together with transcription factors such as bHLHs, MYBs, ERFs and C3Hs. This study indicates the coordinate regulation of bulblet initiation by carbohydrate metabolism and auxin signaling, serving as a basis for further studies on the molecular mechanism of bulblet initiation in lily and other bulbous flowers. [ABSTRACT FROM AUTHOR]

In this study, we characterized the gene expression profile in the roots of Populus ussuriensis at 0, 6, 12, 24, 48 and 120 h after the start of polyethylene glycol (PEG)-induced drought stress using PacBio single-molecule real-time sequencing (SMRT-seq) and Illumina RNA sequencing. Compared to the control, 2244 differentially expressed genes (DEGs) were identified, and many of these DEGs were associated with the signal transduction, antioxidant system, ion accumulation and drought-inducing proteins. Changes in certain physiological and biochemical indexes, such as antioxidant activity and the contents of Ca2+, proline, and total soluble sugars, were further confirmed in P. ussuriensis roots. Furthermore, most of the differentially expressed transcription factors were members of the AP2/ERF, C2H2, MYB, NAC, C2C2 and WRKY families. Additionally, based on PacBio SMRT-seq results, 5955 long non-coding RNAs and 700 alternative splicing events were identified. Our results provide a global view of the gene expression profile that contributes to drought resistance in P. ussuriensis and meaningful information for genetic engineering research in the future. [ABSTRACT FROM AUTHOR]

Populus ussuriensis is an important and fast-growing afforestation plant species in north-eastern China. The whole-genome sequencing of P. ussuriensis has not been completed. Also, the transcriptional network of P. ussuriensis response to cold stress remains unknown. To unravel the early response of P. ussuriensis to chilling (3 °C) stress and freezing (−3 °C) stresses at the transcriptional level, we performed single-molecule real-time (SMRT) and Illumina RNA sequencing for P. ussuriensis. The SMRT long-read isoform sequencing led to the identification of 29,243,277 subreads and 575,481 circular consensus sequencing reads. Approximately 50,910 high-quality isoforms were generated, and 2272 simple sequence repeats and 8086 long non-coding RNAs were identified. The Ca2+ content and abscisic acid (ABA) content in P. ussuriensis were significantly increased under cold stresses, while the value in the freezing stress treatment group was significantly higher than the chilling stress treatment group. A total of 49 genes that are involved in the signal transduction pathways related to perception and transmission of cold stress signals, such as the Ca2+ signaling pathway, ABA signaling pathway and MAPK signaling cascade, were found to be differentially expressed. In addition, 158 transcription factors from 21 different families, such as MYB, WRKY and AP2/ERF, were differentially expressed during chilling and freezing treatments. Moreover, the measurement of physiological indicators and bioinformatics observations demonstrated the altered expression pattern of genes involved in reactive oxygen species balance and the sugar metabolism pathway during chilling and freezing stresses. This is the first report of the early responses of P. ussuriensis to cold stress, which lays the foundation for future studies on the regulatory mechanisms in cold-stress response. In addition the full-length reference transcriptome of P. ussuriensis deciphered could be used in future studies on P. ussuriensis. [ABSTRACT FROM AUTHOR]

Background: Variation in locomotor capacity among animals often reflects adaptations to different environments. Despite evidence that physical performance is heritable, the molecular basis of locomotor performance and performance trade-offs remains poorly understood. In this study we identify the genes, signaling pathways, and regulatory processes possibly responsible for the trade-off between burst performance and endurance observed in Xenopus allofraseri, using a transcriptomic approach., Results: We obtained a total of about 121 million paired-end reads from Illumina RNA sequencing and analyzed 218,541 transcripts obtained from a de novo assembly. We identified 109 transcripts with a significant differential expression between endurant and burst performant individuals (FDR ≤ 0.05 and logFC ≥2), and blast searches resulted in 103 protein-coding genes. We found major differences between endurant and burst-performant individuals in the expression of genes involved in the polymerization and ATPase activity of actin filaments, cellular trafficking, proteoglycans and extracellular proteins secreted, lipid metabolism, mitochondrial activity and regulators of signaling cascades. Remarkably, we revealed transcript isoforms of key genes with functions in metabolism, apoptosis, nuclear export and as a transcriptional corepressor, expressed in either burst-performant or endurant individuals. Lastly, we find two up-regulated transcripts in burst-performant individuals that correspond to the expression of myosin-binding protein C fast-type (mybpc2). This suggests the presence of mybpc2 homoeologs and may have been favored by selection to permit fast and powerful locomotion., Conclusion: These results suggest that the differential expression of genes belonging to the pathways of calcium signaling, endoplasmic reticulum stress responses and striated muscle contraction, in addition to the use of alternative splicing and effectors of cellular activity underlie locomotor performance trade-offs. Ultimately, our transcriptomic analysis offers new perspectives for future analyses of the role of single nucleotide variants, homoeology and alternative splicing in the evolution of locomotor performance trade-offs.

Cephalotaxus oliveri is a tertiary relict conifer endemic to China, regarded as a national second-level protected plant in China. This species has experienced severe changes in temperature and precipitation in the past millions of years, adapting well to harsh environments. In view of global climate change and its endangered conditions, it is crucial to study how it responds to changes in temperature and precipitation for its conservation work. In this study, single-molecule real-time (SMRT) sequencing and Illumina RNA sequencing were combined to generate the complete transcriptome of C. oliveri . Using the RNA-seq data to correct the SMRT sequencing data, the four tissues obtained 63,831 (root), 58,108 (stem), 33,013 (leaf) and 62,436 (male cone) full-length unigenes, with a N50 length of 2523, 3480, 3181, and 3267 bp, respectively. Additionally, 35,887, 11,306, 36,422, and 25,439 SSRs were detected for the male cone, leaf, root, and stem, respectively. The number of long non-coding RNAs predicted from the root was the largest (11,113), and the other tissues were 3408 (stem), 3193 (leaf), and 3107 (male cone), respectively. Functional annotation and enrichment analysis of tissue-specific expressed genes revealed the special roles in response to environmental stress and adaptability in the different four tissues. We also characterized the gene families and pathways related to abiotic factors. This work provides a comprehensive transcriptome resource for C. oliveri , and this resource will facilitate further studies on the functional genomics and adaptive evolution of C. oliveri .

Pseudotaxus chienii , a rare tertiary relict species with economic and ecological value, is a representative of the monotypic genus Pseudotaxus that is endemic to China. P. chienii can adapt well to habitat isolation and ecological heterogeneity under a variety of climate and soil conditions, and is able to survive in harsh environments. However, little is known about the molecular and genetic resources of this long-lived conifer. Herein, we sequenced the transcriptomes of four organs of P. chienii using the PacBio Isoform Sequencing and Illumina RNA Sequencing platforms. Based on the PacBio Iso-Seq data, we obtained 44,896, 58,082, 50,485, and 67,638 full-length unigenes from the root, stem, leaf, and strobilus, respectively, with a mean length of 2692 bp, and a mean N50 length of 3010.75 bp. We then comprehensively annotated these unigenes. The number of organ-specific expressed unigenes ranged from 4393 in leaf to 9124 in strobilus, suggesting their special roles in physiological processes, organ development, and adaptability in the different four organs. A total of 16,562 differentially expressed genes (DEGs) were identified among the four organs and clustered into six subclusters. The gene families related to biotic/abiotic factors, including the TPS, CYP450, and HSP families, were characterized. The expression levels of most DEGs in the phenylpropanoid biosynthesis pathway and plant-pathogen interactions were higher in the root than in the three other organs, suggesting that root constitutes the main organ of defensive compound synthesis and accumulation and has a stronger ability to respond to stress. The sequences were analyzed to predict transcription factors, long non-coding RNAs, and alternative splicing events. The expression levels of most DEGs of C2H2, C3H, bHLH, and bZIP families in the root and stem were higher than those in the leaf and strobilus, indicating that these TFs may play a crucial role in the survival of the root and stem. These results comprise the first comprehensive gene expression profiles obtained for different organs of P. chienii . Our findings will facilitate further studies on the functional genomics, adaptive evolution, and phylogeny of P. chienii , and lay the foundation for the development of conservation strategies for this endangered conifer.

Unilateral eyestalk ablation in the female black tiger shrimp Penaeus monodon is commonly employed to induce ovarian maturation. However, the importance of complementing this practice with the provision of live feed supplement (such as polychaetes) has not been emphasized in previous studies. Indeed, it has been less emphasized that female broodstock must be fed with live feeds such as polychaetes for this practice to be effective. While the effects of eyestalk ablation have been thoroughly studied in various aspects, the synergistic effects of feeding with live feeds and the ablation have never been elucidated at a transcriptome-wide level. With recent advances in the next-generation sequencing platforms, it is now possible to investigate the effects of eyestalk ablation and live feeds at the transcriptomic levels. This study employed both short-read Illumina RNA sequencing and long-read Pacific Biosciences (PacBio) isoform sequencing (Iso-seq) to generate the first high-quality ovarian reference transcriptome in P. monodon. This novel assembly allowed us to dissect the effects of feeds and eyestalk ablation and reveal their synergistic effects at the transcriptomic level through the regulation of important genes involved in fatty acid regulation, energy production, and hormone-mediated oocyte maturation pathways. The synergistic effects between the polychaete feeding and the eyestalk ablation in the process of ovarian maturation in black tiger shrimp suggest that without having proper nutrients from the polychaetes, female broodstock might not be ready to develop its ovary. However, even with proper nutrients, the eyestalk ablation is still necessary to perhaps manipulate the female endocrine of the black tiger shrimp. These findings shed the light on molecular mechanisms and key molecular pathways that lead to successful ovarian maturation.

Nitrogen (N) is the most important limiting factor for cotton production worldwide. Genotype-dependent ability to cope with N shortage has been only partially explored in cotton, and in this context, the comparison of molecular responses of cotton genotypes with different nitrogen use efficiency (NUE) is of particular interest to dissect the key molecular mechanisms underlying NUE. In this study, we employed Illumina RNA-Sequencing to determine the genotypic difference in transcriptome profile using two cotton genotypes differing in NUE (CCRI-69, N-efficient, and XLZ-30, N-inefficient) under N starvation and resupply treatments. The results showed that a large genetic variation existed in differentially expressed genes (DEGs) related to amino acid, carbon, and nitrogen metabolism between CCRI-69 and XLZ-30. Further analysis of metabolic changes in cotton genotypes under N resupply showed that nitrogen metabolism and aromatic amino acid metabolism pathways were mainly enriched in CCRI-69 by regulating carbon metabolism pathways such as starch and sucrose metabolism, glycolysis/gluconeogenesis, and pentose phosphate pathway. Additionally, we performed an expression network analysis of genes related to amino acid, carbon, and nitrogen metabolism. In total, 75 and 33 genes were identified as hub genes in shoots and roots of cotton genotypes, respectively. In summary, the identified hub genes may provide new insights into coordinating carbon and nitrogen metabolism and improving NUE in cotton.

Presently, concern regarding the effects of selenium (Se) on the environment and organisms worldwide is increasing. Too much Se in the soil is harmful to plants. In this study, Illumina RNA sequencing and the untargeted metabolome of control and Se-treated celery seedlings were analyzed. In total, 297,911,046 clean reads were obtained and assembled into 150,218 transcripts (50,876 unigenes). A total of 36,287 unigenes were annotated using different databases. Additionally, 8,907 differentially expressed genes, including 5,319 up- and 3,588 downregulated genes, were identified between mock and Se-treated plants. "Phenylpropanoid biosynthesis" was the most enriched KEGG pathway. A total of 24 sulfur and selenocompound metabolic unigenes were differentially expressed. Furthermore, 1,774 metabolites and 237 significant differentially accumulated metabolites were identified using the untargeted metabolomic approach. We conducted correlation analyses of enriched KEGG pathways of differentially expressed genes and accumulated metabolites. Our findings suggested that candidate genes and metabolites involved in important biological pathways may regulate Se tolerance in celery. The results increase our understanding of the molecular mechanism responsible for celery's adaptation to Se stress., Competing Interests: The authors have declared that no competing interests exist.

Background: Clerodendrum inerme (L.) Gaertn, a halophyte, usually grows on coastal beaches as an important mangrove plant. The salt-tolerant mechanisms and related genes of this species that respond to short-term salinity stress are unknown for us. The de novo transcriptome of C. inerme roots was analyzed using next-generation sequencing technology to identify genes involved in salt tolerance and to better understand the response mechanisms of C. inerme to salt stress., Results: Illumina RNA-sequencing was performed on root samples treated with 400 mM NaCl for 0 h, 6 h, 24 h, and 72 h to investigate changes in C. inerme in response to salt stress. The de novo assembly identified 98,968 unigenes. Among these unigenes, 46,085 unigenes were annotated in the NCBI non-redundant protein sequences (NR) database, 34,756 sequences in the Swiss-Prot database and 43,113 unigenes in the evolutionary genealogy of genes: Non-supervised Orthologous Groups (eggNOG) database. 52 Gene Ontology (GO) terms and 31 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were matched to those unigenes. Most differentially expressed genes (DEGs) related to the GO terms "single-organism process", "membrane" and "catalytic activity" were significantly enriched while numerous DEGs related to the plant hormone signal transduction pathway were also significantly enriched. The detection of relative expression levels of 9 candidate DEGs by qRT-PCR were basically consistent with fold changes in RNA sequencing analysis, demonstrating that transcriptome data can accurately reflect the response of C. inerme roots to salt stress., Conclusions: This work revealed that the response of C. inerme roots to saline condition included significant alteration in response of the genes related to plant hormone signaling. Besides, our findings provide numerous salt-tolerant genes for further research to improve the salt tolerance of functional plants and will enhance research on salt-tolerant mechanisms of halophytes.

Background: Schistosoma japonicum (S. japonicum) is an important zoonotic parasite that is prevalent in China and parts of Southeast Asia. Water buffaloes are an important reservoir and the main transmission sources of S. japonicum. However, self-curing and resistance to re-infection have been observed in water buffaloes., Results: In this study, we compared the morphometry and differences in transcriptional expression of adult S. japonicum worms recovered from primary-infected and re-infected water buffaloes using Illumina RNA-sequencing (RNA-Seq) technology. Results of morphometry analysis revealed that adult S. japonicum worms recovered from re-infected water buffaloes were runtish with smaller organs. The ventral length of male worms was shorter in re-infected buffaloes (328 ± 13 vs 273 ± 8 µm, P < 0.05), and in female worms the oral sucker length (44 ± 3 vs 33 ± 5 µm, P < 0.05), ovary length (578 ± 23 vs 297 ± 27 µm, P < 0.05) and width (150 ± 8 vs 104 ± 9 µm, P < 0.05) were shorter, with fewer eggs in the uteri (41 ± 2 vs 12 ± 1, P < 0.05). Of 13,605 identified genes, 112 were differentially expressed, including 51 upregulated and 61 downregulated genes, in worms from re-infected compared with primary-infected water buffaloes. Gene ontology (GO) enrichment analysis revealed that GO terms such as "oxidation-reduction process", "calcium-dependent phospholipid binding", "lipid binding" and "calcium ion binding" were significantly enriched in downregulated genes, whereas GO terms related to metabolism and biosynthesis were significantly enriched in upregulated genes. The results revealed that the downregulation of some important genes might contribute to a reduction in worm numbers and maldevelopment of surviving worms in re-infected water buffaloes. Furthermore, upregulation of genes related to metabolic processes and biosynthesis might be a compensatory mechanism of worms in disadvantageous environments., Conclusions: To our knowledge, our results present the first large-scale transcriptional expression study identifying the differences between adult S. japonicum worms from primary-infected and re-infected water buffaloes, and particularly emphasize differential expression that may affect the survival and growth of worms in re-infected water buffalo. This will provide new insight into screening for anti-schistosome targets and vaccine candidates.

Conjunctivochalasis (CCH) is a common ocular disease, especially in aged people. However, the molecular mechanism of CCH on transcriptional level has been unclear. In this study, we characterized the transcriptional landscape of human conjunctiva and compared the transcriptome between normal persons (n = 10) and CCH patients (n = 11). Illumina RNA sequencing (RNA-seq) was performed to obtain transcriptional data, and these data were analyzed using various bioinformatics methods, including read mapping, the analysis of gene expression, gene ontology (GO) analysis and Kyoto Encyclopedia of Gene and Genomes (KEGG) metabolic pathway analysis. Additionally, expression patterns of 20 dysregupated genes were validated by qRT-PCR. RNA-seq result showed that clean ratios of 21 samples were more than 95% and more than 92% of all clean reads (32-41 million reads) were mapped to human genome sequence. There were 175 up-regulated genes and 582 down-regulated genes identified in CCH compared to normal persons. Among down-regulated genes in CCH, many genes were related with cell cycle and proliferation, such as BUB1, CCNB1, CCNB2 and CENPA, which might disturb cell growth and proliferation. In addition, several down-regulated genes were associated with keratinization and differentiation of epidermal cells, such as SPRR1A, SPRR1B, and CALML5. In over-expressed genes, CALML6 might play important roles on the development of CCH. The results of qRT-PCR confirmed the accuracy and credibility of RNA-Seq analysis. This study provided a lot of valuable information about pathogenic mechanism of CCH, which could be used to better study CCH in the future., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Despite efforts to restore Atlantic sturgeon in European rivers, aquaculture techniques result in animals with high post-release mortality due to, among other reasons, their low tolerance to increasing water temperature. Marker genes to monitor heat stress are needed in order to identify heat-resistant fish. Therefore, an Atlantic sturgeon cell line was exposed to different heat shock protocols (30 °C and 35 °C) and differences in gene expression were investigated. In total 3020 contigs (∼1.5%) were differentially expressed. As the core of the upregulated contigs corresponded to heat shock proteins (HSP), the heat shock factor (HSF) and the HSP gene families were annotated in Atlantic sturgeon and mapped via Illumina RNA sequencing to identify heat-inducible family members. Up to 6 hsf and 76 hsp genes were identified in the Atlantic sturgeon transcriptome resources, 16 of which were significantly responsive to the applied heat shock. The previously studied hspa1 (hsp70) gene was only significantly upregulated at the highest heat shock (35 °C), while a set of 5 genes (hspc1, hsph3a, hspb1b, hspb11a, and hspb11b) was upregulated at all conditions. Although the hspc1 (hsp90a) gene was previously used as heat shock-marker in sturgeons, we found that hspb11a is the most heat-inducible gene, with up to 3296-fold higher expression in the treated cells, constituting the candidate gene markers for in vivo trials., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Pigs are important, both for agriculture and as animal models for human diseases. However, due to the lack of embryonic stem cells, the possibility of genetic modification is quite limited. To overcome this limitation, induced pluripotent stem (iPS) cells have been derived from pigs. Despite the public availability of a large number of expression datasets from mice, rats, and primates-derived iPS cells, the expression profile of pig-derived iPS cells is quite limited. Furthermore, there is no dataset focused on the profiling of pig-derived iPS cell with six reprogramming factors (Oct3/4, Sox2, Klf4, c-Myc, Lin28, and Nanog). Here, we used Illumina RNA sequencing platform to characterize the mRNA expression of four-factor derived and six-factor derived pig iPS cells. We observed that the expression levels of whole genes in our established six factors derived iPS cells and parent fibroblast, and compared with that of iPS cells with four factors in public database. These data are valuable in understanding species difference in the reprogramming process of stem cells, and could help identify the key regulating genes involved in the process.

Background: Wucai (Brassica campestris L. ssp. chinensis var. rosularis Tsen) is a variant of nonheading Chinese cabbage (Brassica campestris L.), which is one of the major vegetables in China. Cytoplasmic male sterility (CMS) has been used for Wucai breeding in recent years. However, the underlying molecular mechanism of Wucai CMS remains unclear. In this study, the phenotypic and cytological features of Wucai CMS were observed by anatomical analysis, and a comparative transcriptome analysis was carried out to identify genes related to male sterility using Illumina RNA sequencing technology (RNA-Seq)., Results: Microscopic observation demonstrated that tapetum development was abnormal in the CMS line, which failed to produce fertile pollen. Bioinformatics analysis detected 4430 differentially expressed genes (DEGs) between the fertile and sterile flower buds. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed to better understand the functions of these DEGs. Among the DEGs, 35 genes (53 DEGS) were implicated in anther and pollen development, and 11 genes were involved in pollen cell wall formation and modification; most of these showed downregulated expression in sterile buds. In addition, several genes related to tapetum development (A6, AMS, MS1, MYB39, and TSM1) and a few genes annotated to flowering (CO, AP3, VIN3, FLC, FT, and AGL) were detected and confirmed by qRT-PCR as being expressed at the meiosis, tetrad, and uninucleate microspore stages, thus implying possible roles in specifying or determining the fate and development of the tapetum, male gametophyte and stamen. Moreover, the top four largest transcription factor families (MYB, bHLH, NAC and WRKY) were analyzed, and most showed reduced expression in sterile buds. These differentially expressed transcription factors might result in abortion of pollen development in Wucai., Conclusion: The present comparative transcriptome analysis suggested that many key genes and transcription factors involved in anther development show reduced gene expression patterns in the CMS line, which might contribute to male sterility in Wucai. This study provides valuable information for a better understanding of CMS molecular mechanisms and functional genome studies in Wucai.

Transcriptome profiling has become routine in studies of many biological processes. However, the favored approaches such as short-read Illumina RNA sequencing are giving way to long-read sequencing platforms better suited to interrogating the complex transcriptomes typical of many RNA and DNA viruses. Here, we provide a guide-tailored to molecular virologists-to the ins and outs of viral transcriptome sequencing and discuss the strengths and weaknesses of the major RNA sequencing technologies as tools to analyze the abundance and diversity of the viral transcripts made during infection., (Copyright © 2018 American Society for Microbiology.)

Genomic rearrangements that give rise to oncogenic gene fusions can offer actionable targets for cancer therapy. Here we present a systematic analysis of oncogenic gene fusions among a clinically well-characterized, prospectively collected set of 278 primary colon cancers spanning diverse tumor stages and clinical outcomes. Gene fusions and somatic genetic variations were identified in fresh frozen clinical specimens by Illumina RNA-sequencing, the STAR fusion gene detection pipeline, and GATK RNA-seq variant calling. We considered gene fusions to be pathogenically relevant when recurrent, producing divergent gene expression (outlier analysis), or as functionally important (e.g., kinase fusions). Overall, 2.5% of all specimens were defined as harboring a relevant gene fusion (kinase fusions 1.8%). Novel configurations of BRAF, NTRK3 , and RET gene fusions resulting from chromosomal translocations were identified. An R-spondin fusion was found in only one tumor (0.35%), much less than an earlier reported frequency of 10% in colorectal cancers. We also found a novel fusion involving USP9X-ERAS formed by chromothripsis and leading to high expression of ERAS, a constitutively active RAS protein normally expressed only in embryonic stem cells. This USP9X-ERAS fusion appeared highly oncogenic on the basis of its ability to activate AKT signaling. Oncogenic fusions were identified only in lymph node-negative tumors that lacked BRAF or KRAS mutations. In summary, we identified several novel oncogenic gene fusions in colorectal cancer that may drive malignant development and offer new targets for personalized therapy. Cancer Res; 77(14); 3814-22. ©2017 AACR ., (©2017 American Association for Cancer Research.)

Background: Quantitative trait loci (QTLs) detected in one mapping population may not be detected in other mapping populations at all the time. Therefore, before being used for marker assisted breeding, QTLs need to be validated in different environments and/or genetic backgrounds to rule out statistical anomalies. In this regard, we mapped the QTLs controlling various agronomic traits in a recombinant inbred line (RIL) population in response to Nitrogen (N) stress and validated these with the reported QTLs in our earlier study to find the stable and consistent QTLs across populations. Also, with Illumina RNA-sequencing we checked the differential expression of gene (DEG) transcripts between parents and pools of RILs with high and low nitrogen use efficiency (NUE) and overlaid these DEGs on to the common validated QTLs to find candidate genes associated with N-stress tolerance in sorghum., Results: An F 7 RIL population derived from a cross between CK60 (N-stress sensitive) and San Chi San (N-stress tolerant) inbred sorghum lines was used to map QTLs for 11 agronomic traits tested under different N-levels. Composite interval mapping analysis detected a total of 32 QTLs for 11 agronomic traits. Validation of these QTLs revealed that of the detected, nine QTLs from this population were consistent with the reported QTLs in earlier study using CK60/China17 RIL population. The validated QTLs were located on chromosomes 1, 6, 7, 8, and 9. In addition, root transcriptomic profiling detected 55 and 20 differentially expressed gene (DEG) transcripts between parents and pools of RILs with high and low NUE respectively. Also, overlay of these DEG transcripts on to the validated QTLs found candidate genes transcripts for NUE and also showed the expected differential expression. For example, DEG transcripts encoding Lysine histidine transporter 1 (LHT1) had abundant expression in San Chi San and the tolerant RIL pool, whereas DEG transcripts encoding seed storage albumin, transcription factor IIIC (TFIIIC) and dwarfing gene (DW2) encoding multidrug resistance-associated protein-9 homolog showed abundant expression in CK60 parent, similar to earlier study., Conclusions: The validated QTLs among different mapping populations would be the most reliable and stable QTLs across germplasm. The DEG transcripts found in the validated QTL regions will serve as future candidate genes for enhancing NUE in sorghum using molecular approaches.

Background: Despite the economic importance of sugarcane in sugar and bioenergy production, there is not yet a reference genome available. Most of the sugarcane transcriptomic studies have been based on Saccharum officinarum gene indices (SoGI), expressed sequence tags (ESTs) and de novo assembled transcript contigs from short-reads; hence knowledge of the sugarcane transcriptome is limited in relation to transcript length and number of transcript isoforms., Results: The sugarcane transcriptome was sequenced using PacBio isoform sequencing (Iso-Seq) of a pooled RNA sample derived from leaf, internode and root tissues, of different developmental stages, from 22 varieties, to explore the potential for capturing full-length transcript isoforms. A total of 107,598 unique transcript isoforms were obtained, representing about 71% of the total number of predicted sugarcane genes. The majority of this dataset (92%) matched the plant protein database, while just over 2% was novel transcripts, and over 2% was putative long non-coding RNAs. About 56% and 23% of total sequences were annotated against the gene ontology and KEGG pathway databases, respectively. Comparison with de novo contigs from Illumina RNA-Sequencing (RNA-Seq) of the internode samples from the same experiment and public databases showed that the Iso-Seq method recovered more full-length transcript isoforms, had a higher N50 and average length of largest 1,000 proteins; whereas a greater representation of the gene content and RNA diversity was captured in RNA-Seq. Only 62% of PacBio transcript isoforms matched 67% of de novo contigs, while the non-matched proportions were attributed to the inclusion of leaf/root tissues and the normalization in PacBio, and the representation of more gene content and RNA classes in the de novo assembly, respectively. About 69% of PacBio transcript isoforms and 41% of de novo contigs aligned with the sorghum genome, indicating the high conservation of orthologs in the genic regions of the two genomes., Conclusions: The transcriptome dataset should contribute to improved sugarcane gene models and sugarcane protein predictions; and will serve as a reference database for analysis of transcript expression in sugarcane.

Backgroud: Encephalomyocarditis virus (EMCV) has been discovered on pig farms worldwide and can cause myocarditis in piglets and reproductive failure in sows. However, little is known about the host transcriptional responses to infection and host-pathogen interactions., Methods: In this study, transcription profiling was performed by Illumina RNA-Sequencing (RNA-seq) to identify EMCV induced differentially expressed genes in BHK-21 cells at serial time points (12, 24, and 30 h post infection (hpi)), using mock infected cells as control., Results: We identified 237, 241, and 207 differentially expressed genes (DEGs) respectively, majority of which were up-regulated. A large number of DEGs clustered into host defense, cellular signaling and metabolism categories. Moreover, short time series expression analysis revealed that 12 hpi was an important time point for expression change, indicating host virus resistance., Conclusions: This RNA-seq analysis provides the first data for understanding the network of virus host interactions under EMCV infection in vitro, and for identifying host components which involved in the virus infection course.

Haloxylon ammodendron and Haloxylon persicum are important droughttolerant plants in northwest China. The whole-genome sequencing of H. ammodendron and H. persicum grown in their natural environment is incomplete, and their transcriptional regulatory network in response to drought environment remains unclear. To reveal the transcriptional responses of H. ammodendron and H. persicum to an arid environment, we performed single-molecule real-time (SMRT) and Illumina RNA sequencing. In total, 20,246,576 and 908,053 subreads and 435,938 and 210,334 circular consensus sequencing (CCS) reads were identified by SMRT sequencing of H. ammodendron and H. persicum, and 15,238 and 10,135 unigenes, respectively, were successfully obtained. In addition, 9,794 and 7,330 simple sequence repeats (SSRs) and 838 and 71 long non-coding RNAs were identified. In an arid environment, the growth of H. ammodendron was restricted; plant height decreased significantly; basal and branch diameters became thinner and hydrogen peroxide (H2O2) content and peroxidase (POD) activity were increased. Under dry and wet conditions, 11,803 and 15,217 differentially expressed genes (DEGs) were identified in H. ammodendron and H. persicum, respectively. There were 319 and 415 DEGs in the signal transduction pathways related to drought stress signal perception and transmission, including the Ca2C signal pathway, the ABA signal pathway, and the MAPK signal cascade. In addition, 217 transcription factors (TFs) and 398 TFs of H. ammodendron and H. persicum were differentially expressed, including FAR1, MYB, and AP2/ERF. Bioinformatic analysis showed that under drought stress, the expression patterns of genes related to active oxygen [reactive oxygen species (ROS)] scavenging, functional proteins, lignin biosynthesis, and glucose metabolism pathways were altered. Thisis the first full-length transcriptome report concerning the responses of H. ammodendron and H. persicum to drought stress. The results provide a foundation for further study of the adaptation to drought stress. The full-length transcriptome can be used in genetic engineering research. [ABSTRACT FROM AUTHOR]

In this paper, we present a comparative analysis of the transcriptomic profile of three different human cell types: hematopoietic stem cells (HSCs), bone marrow-derived mesenchymal stem cells (MSCs) and fibroblasts (FIBs). The work aims to identify unique genes that are differentially expressed as specific markers of bone marrow-derived MSCs, and to achieve this undertakes a detailed analysis of three independent datasets that include quantification of the global gene expression profiles of three primary cell types: HSCs, MSCs and FIBs. A robust bioinformatics method, called GlobalTest, is used to assess the specific association between one or more genes expressed in a sample and the outcome variable, that is, the 'cell type' provided as a single univariate response. This outcome variable is predicted for each sample tested, based on the expression profile of the specific genes that are used as input to the test. The precision of the tests is calculated along with the statistical sensitivity and specificity for each gene in each dataset, yielding four genes that mark MSCs with high accuracy. Among these, the best performer is the protein-coding gene Transgelin (TAGLN, Gene ID: 6876) (with a Positive Predictive Value > 0.96 and FDR < 0.001), which identifies MSCs better than any of the currently used standard markers: ENG (CD105), THY1 (CD90) or NT5E (CD73). The results are validated by RT-qPCR, providing novel gene biomarkers specific for human MSCs. [ABSTRACT FROM AUTHOR]

Osmotic stress adversely affects the growth, fruit quality and yield of watermelon (Citrullus lanatus (Thunb.) Matsum. & Nakai). Increasing the tolerance of watermelon to osmotic stress caused by factors such as high salt and water deficit is an effective way to improve crop survival in osmotic stress environments. Roots are important organs in water absorption and are involved in the initial response to osmosis stress; however, few studies have examined the underlying mechanism of tolerance to osmotic stress in watermelon roots. For better understanding of this mechanism, the inbred watermelon accession M08, which exhibits relatively high tolerance to water deficits, was treated with 20% polyethylene glycol (PEG) 6000. The root samples were harvested at 6 h after PEG treatment and untreated samples were used as controls. Transcriptome analyses were carried out by Illumina RNA sequencing. A total of 5246 differentially expressed genes were identified. Gene ontology enrichment and biochemical pathway analyses of these 5246 genes showed that short-term osmotic stress affected osmotic adjustment, signal transduction, hormone responses, cell division, cell cycle and ribosome, and M08 may repress root growth to adapt osmotic stress. The results of this study describe the watermelon root transcriptome under osmotic stress and propose new insight into watermelon root responses to osmotic stress at the transcriptome level. Accordingly, these results allow us to better understand the molecular mechanisms of watermelon in response to drought stress and will facilitate watermelon breeding projects to improve drought tolerance., Competing Interests: The authors have declared that no competing interests exist.

Background: Schistosoma turkestanicum is a parasite of considerable veterinary importance as an agent of animal schistosomiasis in many countries, including China. The S. turkestanicum cercariae can also infect humans, causing cercarial dermatitis in many countries and regions of the world. In spite of its significance as a pathogen of animals and humans, there is little transcriptomic and genomic data in the public databases., Methods: Herein, we performed the transcriptome Illumina RNA sequencing (RNA-seq) of adult males and females of S. turkestanicum and de novo transcriptome assembly., Results: Approximately 81.1 (female) and 80.5 (male) million high-quality clean reads were obtained and then 29,526 (female) and 41,346 (male) unigenes were assembled. A total of 34,624 unigenes were produced from S. turkestanicum females and males, with an average length of 878 nucleotides (nt) and N50 of 1480 nt. Of these unigenes, 25,158 (72.7 %) were annotated by blast searches against the NCBI non-redundant protein database. Among these, 21,995 (63.5 %), 22,189 (64.1 %) and 13,754 (39.7 %) of the unigenes had significant similarity in the NCBI non-redundant protein (NR), non-redundant nucleotide (NT) and Swiss-Prot databases, respectively. In addition, 3150 unigenes were identified to be expressed specifically in females and 1014 unigenes were identified to be expressed specifically in males. Interestingly, several pathways associated with gonadal development and sex maintenance were found, including the Wnt signaling pathway (103; 2 %) and progesterone-mediated oocyte maturation (77; 1.5 %)., Conclusions: The present study characterized and compared the transcriptomes of adult female and male blood fluke, S. turkestanicum. These results will not only serve as valuable resources for future functional genomics studies to understand the molecular aspects of S. turkestanicum, but also will provide essential information for ongoing whole genome sequencing efforts on this pathogenic blood fluke.

Litopenaeus vannamei, one of the most important euryhaline crustaceans, is cultured in seawater, brackish water, and freshwater worldwide. We performed Illumina RNA sequencing of L. vannamei gills, generating 124,914,870; 119,250,450; and 105,487,350 raw reads from the shrimps cultured in seawater, brackish water, and freshwater, respectively. From these reads, 466,293 transcripts were de novo assembled and annotated. Comparative genomic analysis showed that 1752 genes were significantly differentially expressed in the freshwater group compared with the seawater group, including 1242 upregulated and 510 downregulated genes. In addition, 1246 genes were differentially expressed in the brackish group vs. the seawater water group, including 659 upregulated and 587 downregulated genes. These differentially expressed genes were mainly involved in energy metabolism, substance metabolism, ion transport and signal transduction, and genetic process. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis were used to analyze the functional significance of the differentially expressed genes, included those responding to salinity through diverse biological functions and processes and numerous potential genes associated with the osmotic response. L. vannamei responses to the three cultivated salinities were analyzed using next-generation sequencing. The transcriptional database established from the current research adds to the information available on L. vannamei and the findings expand our knowledge of the molecular basis of osmoregulation mechanisms in this species., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Background: Filarial nematodes cause serious and debilitating infections in human populations of tropical countries, contributing to an entrenched cycle of poverty. Only one human filarial parasite, Brugia malayi, can be maintained in rodents in the laboratory setting. It has been a widely used model organism in experiments that employ culture systems, the impact of which on the worms is unknown., Methodology/principal Findings: Using Illumina RNA sequencing, we characterized changes in gene expression upon in vitro maintenance of adult B. malayi female worms at four time points: immediately upon removal from the host, immediately after receipt following shipment, and after 48 h and 5 days in liquid culture media. The dramatic environmental change and the 24 h time lapse between removal from the host and establishment in culture caused a globally dysregulated gene expression profile. We found a maximum of 562 differentially expressed genes based on pairwise comparison between time points. After an initial shock upon removal from the host and shipping, a few stress fingerprints remained after 48 h in culture and until the experiment was stopped. This was best illustrated by a strong and persistent up-regulation of several genes encoding cuticle collagens, as well as serpins., Conclusions/significance: These findings suggest that B. malayi can be maintained in culture as a valid system for pharmacological and biological studies, at least for several days after removal from the host and adaptation to the new environment. However, genes encoding several stress indicators remained dysregulated until the experiment was stopped.

Orchardgrass (Dactylis glomerata L.) is a long-lived, cool-season forage grass that is commonly used for hay production. Despite its economic importance, orchardgrass genome remains relatively unexplored. In this study, we used Illumina RNA sequencing to identify gene-associated molecular markers, including simple sequence repeats (SSRs) and single nucleotide polymorphisms (SNPs), as well as heat stress-induced differentially expressed genes (DEGs) in two orchardgrass genotypes, 'Baoxing' (heat resistant) and '01998' (heat susceptible). Approximately 163 million high-quality trimmed reads were generated from 207 million raw reads using the Illumina HiSeq 2000 platform. A total of 126,846 unigenes were obtained after de novo assembly of the trimmed reads, and 40,078 unigenes were identified as coding sequences (CDSs). Based on the assembled unigenes, 669,300 high-quality SNPs, including 416,099 transitions and 257,736 transversions, were contained in 75,875 unigenes. In addition, a total of 8475 microsatellites were detected in 7764 unigenes. When placed under heat stress, the total number of DEGs in 'Baoxing' (3527) was higher than in '01998' (2649), indicating that in comparison with heat-susceptible '01998', heat-resistant 'Baoxing' seems to have more unigenes that respond to heat stress. The high-throughput transcriptome sequencing of orchardgrass under heat stress provides useful information for gene identification and for the development of SNP and SSR molecular markers. The comparison of DEGs under different periods of heat stress allowed us to identify a wealth of candidate DEGs that can be further analysed in order to determine the genetic mechanisms underlying heat tolerance in orchardgrass., (© 2015 John Wiley & Sons Ltd.)

Current genomic studies are limited by the availability of fresh tissue samples. Here, we show that Illumina RNA sequencing of formalin-fixed diagnostic tumor samples produces gene expression that is strongly correlated with matched frozen tumor samples (r > 0.89). In addition, sequence variations identified from FFPE RNA show 99.67% concordance with that from exome sequencing of matched frozen tumor samples. Because FFPE is a routine diagnostic sample preparation, the feasibility results reported here will facilitate the setup of large-scale research and clinical studies in medical genomics that are currently limited by the availability of fresh frozen samples.

Paeonia lactiflora is one of the most famous species of herbaceous peonies with gorgeous flowers. Bud dormancy is a crucial developmental process that allows P. lactiflora to survive unfavorable environmental conditions. However, little information is available on the molecular mechanism of the bud dormancy in P. lactiflora. We performed de novo transcriptome sequencing using the Illumina RNA sequencing platform for the underground renewal buds of P. lactiflora 'Hangbaishao' to study the molecular mechanism underlying its bud dormancy transition (the period from endodormancy to ecodormancy) and release (the period from ecodormancy to bud elongation and sprouting). Approximately 300 million high-quality clean reads were generated and assembled into 207,827 (mean length = 828 bp) and 51,481 (mean length = 1250 bp) unigenes using two assembly methods named "Trinity" and "Trinity+PRICE", respectively. Based on the data obtained by the latter method, 32,316 unigenes were annotated by BLAST against various databases. Approximately 1,251 putative transcription factors were obtained, of which the largest number of unique transcripts belonged to the basic helix-loop-helix protein (bHLH) transcription factor family, and five of the top ten highly expressed transcripts were annotated as dehydrin (DHN). A total of 17,705 simple sequence repeat (SSR) motifs distributed in 13,797 sequences were obtained. The budbreak morphology, levels of indole-3-acetic acid (IAA) and abscisic acid (ABA), and activities of guaiacol peroxidase (POD) and catalase (CAT) were observed. The expression of 20 interested unigenes, which annotated as DHN, heat shock protein (HSP), histone, late elongated hypocotyl (LHY), and phytochrome (PHY), and so on, were also analyzed. These studies were based on morphological, physiological, biochemical, and molecular levels and provide comprehensive insight into the mechanism of dormancy transition and release in P. lactiflora. Transcriptome dataset can be highly valuable for future investigation on gene expression networks in P. lactiflora as well as research on dormancy in other non-model perennial horticultural crops of commercial significance.

Platycladus orientalis (P. orientalis) is a common tree used for vegetation restoration in northern China, and its large area propagation helps to improve site conditions. However, under harsh conditions such as poor land, the survival rate of P. orientalis is very low. Numerous studies have shown that root pruning can promote the formation of lateral roots in seedlings, enhancing the roots' capacity to absorb soil nutrients and water, and thereby improving the survival rate of seedlings. In this study, a one-third root pruning treatment was applied to P. orientalis seedlings, and the whole transcriptome of seedlings subjected to both control (CK) and root pruning treatments was sequenced to analyze their gene expression profiles. This study investigated the regulatory mechanisms of lateral root development in response to root pruning damage at the molecular level. Using nine cells, 15.28 Gb of clean data were obtained, which yielded 101,688 high-quality full-length transcript sequences and 22,955 low-quality full-length transcript sequences after clustering. Redundancy was then removed using CD-HIT, and Illumina RNA-seq sequencing produced 139.26 Gb of clean data. A total of 2025 differentially expressed genes (DEGs) were identified at three time points following root pruning treatment. Enrichment analysis revealed that the peroxidase gene family plays a significant role in lateral root proliferation. Furthermore, the expression levels of the peroxidase gene family were notably upregulated in comparison to the control group. Pathway enrichment analysis identified 22 relevant genes, which appeared to be highly associated with root growth and resilience to stress. Through examining the expression patterns and correlations of these genes, five central genes emerged as key players. The findings of this research suggest that the peroxidase gene family plays a crucial role in the stress response and root development of P. orientalis, providing reference and guidance for root development in other plant species. [ABSTRACT FROM AUTHOR]

Jerusalem artichoke (Helianthus tuberosus L.) has long been cultivated as a vegetable and as a source of fructans (inulin) for pharmaceutical applications in diabetes and obesity prevention. However, transcriptomic and genomic data for Jerusalem artichoke remain scarce. In this study, Illumina RNA sequencing (RNA-Seq) was performed on samples from Jerusalem artichoke leaves, roots, stems and two different tuber tissues (early and late tuber development). Data were used for de novo assembly and characterization of the transcriptome. In total 206,215,632 paired-end reads were generated. These were assembled into 66,322 loci with 272,548 transcripts. Loci were annotated by querying against the NCBI non-redundant, Phytozome and UniProt databases, and 40,215 loci were homologous to existing database sequences. Gene Ontology terms were assigned to 19,848 loci, 15,434 loci were matched to 25 Clusters of Eukaryotic Orthologous Groups classifications, and 11,844 loci were classified into 142 Kyoto Encyclopedia of Genes and Genomes pathways. The assembled loci also contained 10,778 potential simple sequence repeats. The newly assembled transcriptome was used to identify loci with tissue-specific differential expression patterns. In total, 670 loci exhibited tissue-specific expression, and a subset of these were confirmed using RT-PCR and qRT-PCR. Gene expression related to inulin biosynthesis in tuber tissue was also investigated. Exsiting genetic and genomic data for H. tuberosus are scarce. The sequence resources developed in this study will enable the analysis of thousands of transcripts and will thus accelerate marker-assisted breeding studies and studies of inulin biosynthesis in Jerusalem artichoke.

The marine medaka (Oryzias melastigma) has been increasingly used as a fish model for detecting environmental stresses and chemical contaminants in the marine environment. Recent mammalian studies have shown that environmental stresses can alter the expression profiles of microRNAs (miRNAs), leading to transgenerational effects. Here, we use high-throughput Illumina RNA sequencing (RNA-Seq) for miRNA transcriptome analysis of brain, liver, and gonads from sexually mature male and female marine medaka. A total of 128,883,806 filtered sequence reads were generated from six small RNA libraries, identifying a total of 2,125,663 non-redundant sequences. These sequences were aligned and annotated to known animal miRNAs (miRBase) using the BLAST method. A total of 223 distinct miRNA types were identified, with the greatest number expressed in brain tissue. Our data suggested that 55 miRNA types from 34 families are common to all tested tissues, while some of the miRNAs are tissue-enriched or sex-enriched. Quantitative real-time PCR analysis further demonstrated that let-7a, miR-122, and miR-9-3p were downregulated in hypoxic female medaka, while miR-2184 was specifically upregulated in the testis of hypoxic male fish. This is the first study to identify miRNAs in O. melastigma using small RNA deep sequencing technology. Because miRNA expression is highly conserved between marine medaka and other vertebrates, marine medaka may serve as a good model for studies on the functional roles of miRNAs in hypoxia stress response and signaling in marine fish.

Background: There is growing interest in odonates (damselflies and dragonflies) as model organisms in ecology and evolutionary biology but the development of genomic resources has been slow. So far only one draft genome (Ladona fulva) and one transcriptome assembly (Enallagma hageni) have been published. Odonates have some of the most advanced visual systems among insects and several species are colour polymorphic, and genomic and transcriptomic data would allow studying the genomic architecture of these interesting traits and make detailed comparative studies between related species possible. Here, we present a comprehensive de novo transcriptome assembly for the blue-tailed damselfly Ischnura elegans (Odonata: Coenagrionidae) built from short-read RNA-seq data. The transcriptome analysis in this paper provides a first step towards identifying genes and pathways underlying the visual and colour systems in this insect group., Results: Illumina RNA sequencing performed on tissues from the head, thorax and abdomen generated 428,744,100 paired-ends reads amounting to 110 Gb of sequence data, which was assembled de novo with Trinity. A transcriptome was produced after filtering and quality checking yielding a final set of 60,232 high quality transcripts for analysis. CEGMA software identified 247 out of 248 ultra-conserved core proteins as 'complete' in the transcriptome assembly, yielding a completeness of 99.6%. BLASTX and InterProScan annotated 55% of the assembled transcripts and showed that the three tissue types differed both qualitatively and quantitatively in I. elegans. Differential expression identified 8,625 transcripts to be differentially expressed in head, thorax and abdomen. Targeted analyses of vision and colour functional pathways identified the presence of four different opsin types and three pigmentation pathways. We also identified transcripts involved in temperature sensitivity, thermoregulation and olfaction. All these traits and their associated transcripts are of considerable ecological and evolutionary interest for this and other insect orders., Conclusions: Our work presents a comprehensive transcriptome resource for the ancient insect order Odonata and provides insight into their biology and physiology. The transcriptomic resource can provide a foundation for future investigations into this diverse group, including the evolution of colour, vision, olfaction and thermal adaptation.

Potassium (K) deficiency is one of the major factors affecting crop growth and productivity. Development of low-K tolerant crops is an effective approach to solve the nutritional deficiency in agricultural production. Tibetan annual wild barley is rich in genetic diversity and can grow normally under poor soils, including low-K supply. However, the molecular mechanism about low K tolerance is still poorly understood. In this study, Illumina RNA-Sequencing was performed using two Tibetan wild barley genotypes differing in low K tolerance (XZ153, tolerant and XZ141, sensitive), to determine the genotypic difference in transcriptome profiling. We identified a total of 692 differentially expressed genes (DEGs) in two genotypes at 6 h and 48 h after low-K treatment, including transcription factors, transporters and kinases, oxidative stress and hormone signaling related genes. Meanwhile, 294 low-K tolerant associated DEGs were assigned to transporter and antioxidant activities, stimulus response, and other gene ontology (GO), which were mainly involved in starch and sucrose metabolism, lipid metabolism and ethylene biosynthesis. Finally, a hypothetical model of low-K tolerance mechanism in XZ153 was presented. It may be concluded that wild barley accession XZ153 has a higher capability of K absorption and use efficiency than XZ141 under low K stress. A rapid response to low K stress in XZ153 is attributed to its more K uptake and accumulation in plants, resulting in higher low K tolerance. The ethylene response pathway may account for the genotypic difference in low-K tolerance.

Background: Sorghum is an important cereal crop, which requires large quantities of nitrogen fertilizer for achieving commercial yields. Identification of the genes responsible for low-N tolerance in sorghum will facilitate understanding of the molecular mechanisms of low-N tolerance, and also facilitate the genetic improvement of sorghum through marker-assisted selection or gene transformation. In this study we compared the transcriptomes of root tissues from seven sorghum genotypes having differential response to low-N stress., Results: Illumina RNA-sequencing detected several common differentially expressed genes (DEGs) between four low-N tolerant sorghum genotypes (San Chi San, China17, KS78 and high-NUE bulk) and three sensitive genotypes (CK60, BTx623 and low-NUE bulk). In sensitive genotypes, N-stress increased the abundance of DEG transcripts associated with stress responses including oxidative stress and stimuli were abundant. The tolerant genotypes adapt to N deficiency by producing greater root mass for efficient uptake of nutrients. In tolerant genotypes, higher abundance of transcripts related to high affinity nitrate transporters (NRT2.2, NRT2.3, NRT2.5, and NRT2.6) and lysine histidine transporter 1 (LHT1), may suggest an improved uptake efficiency of inorganic and organic forms of nitrogen. Higher abundance of SEC14 cytosolic factor family protein transcript in tolerant genotypes could lead to increased membrane stability and tolerance to N-stress., Conclusions: Comparison of transcriptomes between N-stress tolerant and sensitive genotypes revealed several common DEG transcripts. Some of these DEGs were evaluated further by comparing the transcriptomes of genotypes grown under full N. The DEG transcripts showed higher expression in tolerant genotypes could be used for transgenic over-expression in sensitive genotypes of sorghum and related crops for increased tolerance to N-stress, which results in increased nitrogen use efficiency for sustainable agriculture.

Maize (Zea mays) is an important food and forage crop, as well as an industrial raw material, that plays important roles in agriculture and national economies. Drought stress has negative effects on seed germination and seedling growth, and it decreases crop production. In this study, we selected two maize inbred lines with different drought-tolerance levels: drought-tolerant 287M and drought-sensitive 753F. The physiological results showed that drought stress resulted in a large accumulation of reactive oxygen species (ROS) in maize root cells. However, in 287M, the activity levels of the ROS scavenging enzymes superoxide dismutase and ascorbate peroxidase also increased, resulting in a higher ROS scavenging ability than 753F. We used Illumina RNA sequencing to obtain the gene expression profiles of the two maize inbred lines at the seedling stage in response to drought stress. The transcriptome data were analyzed to reveal the mechanisms underlying the drought tolerance of 287M at the gene regulatory level. The differences in drought tolerance between 287M and 753F may be associated with different ROS scavenging capabilities, signal interaction networks, and some transcription factors. Our results will aid in understanding the molecular mechanisms involved in plant responses to drought stress. • Drought affected maize growth and ROS accumulation. • Drought induced secondary metabolites, ROS scavenging, and hormone-related genes. • Genes involved in ethylene synthesis were down-regulated in drought-sensitive 753F. • Secondary metabolism and transcription factors were up-regulated in tolerant 287M. • 287M and 753F appear to have different drought-stress response networks. [ABSTRACT FROM AUTHOR]

Tomato is a model and economically important crop plant with little information available about gene expression in roots. Currently, there have only been a few studies that examine hormonal responses in tomato roots and none at a genome-wide level. This study examined the transcriptome atlas of tomato root regions (root tip, lateral roots, and whole roots) and the transcriptional regulation of each root region in response to the plant hormones cytokinin and auxin using Illumina RNA sequencing. More than 165 million 1×54 base pair reads were mapped onto the Solanum lycopersicum reference genome and differential expression patterns in each root region in response to each hormone were assessed. Many novel cytokinin- and auxin-induced and -repressed genes were identified as significantly differentially expressed and the expression levels of several were confirmed by qPCR. A number of these regulated genes represent tomato orthologues of cytokinin- or auxin-regulated genes identified in other species, including CKXs, type-A RRs, Aux/IAAs, and ARFs. Additionally, the data confirm some of the hormone regulation studies for recently examined genes in tomato such as SlIAAs and SlGH3s. Moreover, genes expressed abundantly in each root region were identified which provide a spatial distribution of many classes of genes, including plant defence, secondary metabolite production, and general metabolism across the root. Overall this study presents the first global expression patterns of hormone-regulated transcripts in tomato roots, which will be functionally relevant for future studies directed towards tomato root growth and development.

Microplastics are considered an emerging environmental pollutant due to their ubiquitous presence in the environment. However, the potential impact of microplastics on human health warrants further research. Recent studies have reported neurobehavioral and neurotoxic effects in marine and rodent models; however, their impact on the underlying cellular physiology in mammals remains unclear. Herein, we exposed neural stem cells and neural stem cell-derived astrocytes, oligodendrocytes, and neurons to various sizes and concentrations of polystyrene nano- and microplastics. We investigated their cellular uptake, impact on cytotoxicity, and alteration of gene expression through transcriptome profiling. The cell type most affected by decreased viability were astrocytes after 7 days of repeated exposure. Transcriptional analysis showed that 1274 genes were differentially expressed in astrocytes exposed to 500 nm microplastics, but only 531 genes were altered in astrocytes exposed to 50 nm nanoplastics. Both canonical pathway and Kyoto Encyclopedia of Genes and Genomes analysis showed that upregulated pathways were involved in neuroinflammation, innate and adaptive immunity, cell migration, proliferation, extracellular matrix remodeling, and cytoskeleton structures. The downregulated pathways were involved in lipid metabolism, specifically fatty acid oxidation and cholesterol metabolism. Our results show that neural stem cell-derived astrocytes repeatedly exposed to nano- and microplastics for 7 days undergo changes that are hallmarks of astrogliosis. [ABSTRACT FROM AUTHOR]

• A transcriptome of Populus tomentosa during cold stress has been analyzed. • This work identified numerous cold-related genes involved in sugar metabolisms, antioxidant defense system, hormone signal transduction and photosynthesis. • Our study provides valuable information for further studies on cold response mechanisms and genetic improvement of cold tolerance in Populus tomentosa. Low temperature affects the growth, development, and even survival in plants. Populus tomentosa is a major industrial tree species used for timber and pulp manufacturing in northern China. Low temperature is an important stress factor for this species, and restricts its extensive cultivation and distribution. To face this problem, an understanding of the molecular mechanism underlying responses to cold stress in this species is necessary for molecular breeding. In this study, Illumina RNA-sequencing was employed to comprehensively investigate the gene expression profiles of P. tomentosa in response to cold stress. Two cDNA libraries from cold-treated and control samples were constructed and sequenced, generating a total of 94,226 unigenes. Of these, 1978 were differentially expressed, with 1468 upregulated and 510 downregulated. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses identified the relevant biological functions and metabolic pathways of these differentially expressed genes. Based on enrichment results, a diverse range of cold response candidate genes were obtained and mainly involved in sugar metabolism, antioxidant defense system, hormone signal transduction, and photosynthesis, such as genes encoding sucrose synthase, superoxide dismutase, brassinosteroid-signaling kinase, and ferredoxin. Various transcription factors with divergent expression patterns were identified. Finally, RNA-seq data were validated using real-time quantitative polymerase chain reaction analyses. This study provides valuable information for further studies on cold response mechanisms and genetic improvement of cold tolerance in P. tomentosa. [ABSTRACT FROM AUTHOR]

The apicomplexan parasite Theileria annulata is transmitted by Hyalomma ticks and causes an acute lymphoproliferative disease that is invariably lethal in exotic cattle breeds. The unique ability of the schizont stage of T. annulata to transform infected leukocytes to a cancer-like phenotype and the simplicity of culturing and passaging T. annulata-transformed cells in vitro have been explored for live vaccine development by attenuating the transformed cells using lengthy serial propagation in vitro. The empirical in vivo evaluation of attenuation required for each batch of long-term cultured cells is a major constraint since it is resource intensive and raises ethical issues regarding animal welfare. As yet, the molecular mechanisms underlying attenuation are not well understood. Characteristic changes in gene expression brought about by attenuation are likely to aid in the identification of novel biomarkers for attenuation. We set out to undertake a comparative transcriptome analysis of attenuated (passage 296) and virulent (passage 26) bovine leukocytes infected with a Tunisian strain of T. annulata termed Beja. RNA-seq was used to analyse gene expression profiles and the relative expression levels of selected genes were verified by real-time quantitative PCR (RT-qPCR) analysis. Among the 3538 T. annulata genes analysed, 214 were significantly differentially expressed, of which 149 genes were up-regulated and 65 down-regulated. Functional annotation of differentially expressed T. annulata genes revealed four broad categories of metabolic pathways: carbon metabolism, oxidative phosphorylation, protein processing in the endoplasmic reticulum and biosynthesis of secondary metabolites. It is interesting to note that of the top 40 genes that showed altered expression, 13 were predicted to contain a signal peptide and/or at least one transmembrane domain, suggesting possible involvement in host-parasite interaction. Of the 16,514 bovine transcripts, 284 and 277 showed up-regulated and down-regulated expression, respectively. These were assigned to functional categories relevant to cell surface, tissue morphogenesis and regulation of cell adhesion, regulation of leucocyte, lymphocyte and cell activation. The genetic alterations acquired during attenuation that we have catalogued herein, as well as the accompanying in silico functional characterization, do not only improve understanding of the attenuation process, but can also be exploited by studies aimed at identifying attenuation biomarkers across different cell lines focusing on some host and parasite genes that have been highlighted in this study, such as bovine genes (CD69, ZNF618, LPAR3, and APOL3) and parasite genes such as TA03875. [ABSTRACT FROM AUTHOR]

Expressional profiling of the endometrium enables the personalised timing of the window of implantation (WOI). This study presents and evaluates a novel analytical pipeline based on a TAC-seq (Targeted Allele Counting by sequencing) method for endometrial dating. The expressional profiles were clustered, and differential expression analysis was performed on the model development group, using 63 endometrial biopsies spanning over proliferative (PE, n = 18), early-secretory (ESE, n = 18), mid-secretory (MSE, n = 17) and late-secretory (LSE, n = 10) endometrial phases of the natural cycle. A quantitative predictor model was trained on the development group and validated on sequenced samples from healthy women, consisting of 52 paired samples taken from ESE and MSE phases and five LSE phase samples from 31 individuals. Finally, the developed test was applied to 44 MSE phase samples from a study group of patients diagnosed with recurrent implantation failure (RIF). In validation samples (n = 57), we detected displaced WOI in 1.8% of the samples from fertile women. In the RIF study group, we detected a significantly higher proportion of the samples with shifted WOI than in the validation set of samples from fertile women, 15.9% and 1.8% (p = 0.012), respectively. The developed model was evaluated with an average cross-validation accuracy of 98.8% and an accuracy of 98.2% in the validation group. The developed beREADY screening model enables sensitive and dynamic detection of selected transcriptome biomarkers, providing a quantitative and accurate prediction of endometrial receptivity status. [ABSTRACT FROM AUTHOR]

BACKGROUND: Volatile organic compounds are critical for food flavor and play important roles in plant–plant interactions and plants' communications with the environment. Tobacco is well‐studied for secondary metabolism and most of the typical flavor substances in tobacco leaves are generated at the mature stage of leaf development. However, the changes in volatiles during leaf senescence are rarely studied. RESULTS: The volatile composition of tobacco leaves at different stages of senescence was characterized for the first time. Comparative volatile profiling of tobacco leaves at different stages was performed using solid‐phase microextraction coupled with gas chromatography/mass spectrometry. In total, 45 volatile compounds were identified and quantified, including terpenoids, green leaf volatiles (GLVs), phenylpropanoids, Maillard reaction products, esters, and alkanes. Most of the volatile compounds showed differential accumulation during leaf senescence. Some terpenoids, including neophytadiene, β‐springene, and 6‐methyl‐5‐hepten‐2‐one, increased significantly with the progress of leaf senescence. Hexanal and phenylacetaldehyde also showed increased accumulation in leaves during senescence. The results from gene expression profiling indicated that genes involved in metabolism of terpenoids, phenylpropanoids, and GLVs were differentially expressed during leaf yellowing. CONCLUSION: Dynamic changes in volatile compounds during tobacco leaf senescence are observed and the integration of gene‐metabolites datasets offers important readouts for the genetic control of volatile production during the process of leaf senescence. © 2023 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

Asparagus racemosus is known for its diverse content of secondary metabolites, i.e., saponins, alkaloids, and a wide range of flavonoids. Flavonoids, including phenols andpolyphenols, haveasignificantrole in plantphysiologyandaresynthesized in several tissues. Despite thediverse role offlavonoids, genetic informationis limited forflavonoid biosynthesis pathways in A. racemosus. The current study explores full-scale functional genomics information of A. racemosus by de novo transcriptome sequencing using Illumina paired-endsequencingtechnologyto elucidate thegenesinvolvedinflavonoid biosynthesis pathways. The de novoassembly of high-quality paired-end reads resulted in ~2.3 million high-quality reads with a pooled transcript of 45,647 comprising ~76Mb transcriptome with a mean length (bp) of 1,674 and N50 of 1,868bp. Furthermore, the coding sequence (CDS) prediction analysis from 45,647 pooled transcripts resulted in 45,444 CDSwith a total length and mean length of 76,398,686 and 1,674, respectively. The Gene Ontology (GO) analysis resulted in a high number of CDSs assigned to 25,342 GO terms, which grouped the predicted CDS into three main domains, i.e., Biological Process (19,550), Molecular Function (19,873), and Cellular Component (14,577). The Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway database was used to categorize 6,353 CDS into 25 distinct biological pathway categories, in which the majority of mapped CDS were shown to be related to translation (645), followed by signal transduction (532), carbohydrate metabolism (524), folding, sorting, and degradation (522). Among these, only ~64 and 14 CDSs were found to be involved in the phenylpropanoid and flavonoid biosynthesis pathways, respectively. Quantitative Real-time PCR was used to check the expression profile of fourteen potential flavonoid biosynthesis pathway genes. The qRT-PCR analysis result matches the transcriptome sequence data validating the Illumina sequence results. Moreover, a large number of genes associated with the flavonoids biosynthesis pathway were found to be upregulated under the induction of methyl jasmonate. The present-day studyontranscriptomesequencedata of A.racemosuscanbeutilizedfor characterizing genesinvolvedinflavonoidbiosynthesispathwaysandforfunctionalgenomicsanalysisin A. racemosus using the reverse genetics approach (CRISPR/Cas9 technology). [ABSTRACT FROM AUTHOR]