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author:("Zhao, wensing")
1.  Characterization and comparative profiling of ovarian microRNAs during ovine anestrus and the breeding season 
BMC Genomics  2014;15(1):899.
Background
Seasonal estrus is a critical limiting factor of animal fecundity, and it involves changes in both ovarian biology and hormone secretion in different seasons. Previous studies indicate that two classes of small RNAs (miRNAs and piRNAs) play important regulatory roles in ovarian biology. To understand the roles of small RNA-mediated post-transcriptional regulation in ovine seasonal estrus, the variation in expression patterns of ovarian small RNAs during anestrus and the breeding season were analyzed using Solexa sequencing technology. In addition, reproductive hormone levels were determined during ovine anestrus and the breeding season.
Results
A total of 483 miRNAs (including 97 known, 369 conserved and 17 predicated novel miRNAs), which belong to 183 different miRNA families, were identified in ovaries of Tan sheep and Small Tail Han (STH) sheep. Compared with the three stages of the breeding season, 25 shared significantly differentially expressed (including 19 up- and six down-regulated) miRNAs were identified in ovine anestrus. KEGG Pathway analysis revealed that the target genes for some of the differentially expressed miRNAs were involved in reproductive hormone related pathways (e.g. steroid biosynthesis, androgen and estrogen metabolism and GnRH signaling pathway) as well as follicular/luteal development related pathways. Moreover, the expression of the differentially expressed miRNAs and most of their target genes were negatively correlated in the above pathways. Furthermore, the levels of estrogen, progesterone and LH in ovine anestrus were significantly lower than those in the breeding season. Combining the results of pathway enrichment analysis, expression of target genes and hormone measurement, we suggest that these differentially expressed miRNAs in anestrus might participate in attenuation of ovarian activity by regulating the above pathways. Besides miRNAs, a large and unexpectedly diverse set of piRNAs were also identified.
Conclusions
The miRNA profiles of ovine ovaries in anestrus were presented for the first time. The identification and characterization of miRNAs that are differentially expressed between ovine anestrus and the breeding season will help understanding of the role of miRNAs in the regulation of seasonal estrus, and provides candidates for determining miRNAs which could be potentially used to regulate ovine seasonal estrus.
Electronic supplementary material
The online version of this article (doi:10.1186/1471-2164-15-899) contains supplementary material, which is available to authorized users.
doi:10.1186/1471-2164-15-899
PMCID: PMC4287553  PMID: 25318541
Sheep; Seasonal estrus; Anestrus; Ovary; miRNA; piRNA
2.  WBSA: Web Service for Bisulfite Sequencing Data Analysis 
PLoS ONE  2014;9(1):e86707.
Whole-Genome Bisulfite Sequencing (WGBS) and genome-wide Reduced Representation Bisulfite Sequencing (RRBS) are widely used to study DNA methylation. However, data analysis is complicated, lengthy, and hampered by a lack of seamless analytical pipelines. To address these issues, we developed a convenient, stable, and efficient web service called Web Service for Bisulfite Sequencing Data Analysis (WBSA) to analyze bisulfate sequencing data. WBSA focuses on not only CpG methylation, which is the most common biochemical modification in eukaryotic DNA, but also non-CG methylation, which have been observed in plants, iPS cells, oocytes, neurons and stem cells of human. WBSA comprises three main modules as follows: WGBS data analysis, RRBS data analysis, and differentially methylated region (DMR) identification. The WGBS and RRBS modules execute read mapping, methylation site identification, annotation, and advanced analysis, whereas the DMR module identifies actual DMRs and annotates their correlations to genes. WBSA can be accessed and used without charge either online or local version. WBSA also includes the executables of the Portable Batch System (PBS) and standalone versions that can be downloaded from the website together with the installation instructions. WBSA is available at no charge for academic users at http://wbsa.big.ac.cn.
doi:10.1371/journal.pone.0086707
PMCID: PMC3907392  PMID: 24497972
3.  Molecular Cloning and Functional Analysis of the Duck TLR4 Gene 
Toll-like receptor 4 (TLR4) recognizes pathogen-associated molecular patterns in some animals and has been shown to be closely associated with several diseases such as tumors, atherosclerosis, and asthma. However, its function in ducks is not clear. Alternative splicing of the TLR4 gene has been identified in pigs, sheep, mice, and other species, but has not yet been reported in the duck. In this study, alternative splicing of the duck TLR4 gene was investigated using reverse transcription-polymerase chain reaction (RT-PCR). Duck TLR4 gene (duTLR4, accession number: KF278109) was found to consist of 3367 nucleotides of coding sequence. An alternative splice form, TLR4-b, was identified and shown by alignment to retain the intron between exons 1 and 2. Real-time quantitative polymerase chain reaction (qPCR) analyses suggested that duTLR4-a (wild-type) mRNA is widely expressed in various healthy tissues, whereas TLR4-b is expressed at only low levels. Following stimulation of normal duck embryo fibroblasts with lipopolysaccharide, the expression of both isoforms initially increased and then decreased. Expression of the wild-type isoform subsequently increased again, while that of the variant remained low. The expression levels of wild-type TLR4 were further analyzed by transient transfection of a pcDNA3.1(+)-TLR4-a overexpression vector into duck embryo fibroblasts. qRT-PCR analyses showed that after stimulation with LPS and poly(I:C) the expression levels of IL-1β, IL6, and MHC II increased with a response-efficacy relationship. Our experimental results indicate that TLR4 plays an important role in resistance to both bacterial and viral infections in the duck.
doi:10.3390/ijms140918615
PMCID: PMC3794798  PMID: 24025421
duck; TLR4; alternative splicing; expression analysis
4.  Transcriptome Profiling of the Goose (Anser cygnoides) Ovaries Identify Laying and Broodiness Phenotypes 
PLoS ONE  2013;8(2):e55496.
Background
The geese have strong broodiness and poor egg performance. These characteristics are the key issues that hinder the goose industry development. Yet little is known about the mechanisms responsible for follicle development due to lack of genomic resources. Hence, studies based on high-throughput sequencing technologies are needed to produce a comprehensive and integrated genomic resource and to better understand the biological mechanisms of goose follicle development.
Methodology/Principal Findings
In this study, we performed de novo transcriptome assembly and gene expression analysis using short-read sequencing technology (Illumina). We obtained 67,315,996 short reads of 100 bp, which were assembled into 130,514 unique sequences by Trinity strategy (mean size = 753bp). Based on BLAST results with known proteins, these analyses identified 52,642 sequences with a cut-off E-value above 10−5. Assembled sequences were annotated with gene descriptions, gene ontology and clusters of orthologous group terms. In addition, we investigated the transcription changes during the goose laying/broodiness period using a tag-based digital gene expression (DGE) system. We obtained a sequencing depth of over 4.2 million tags per sample and identified a large number of genes associated with follicle development and reproductive biology including cholesterol side-chain cleavage enzyme gene and dopamine beta-hydroxylas gene. We confirm the altered expression levels of the two genes using quantitative real-time PCR (qRT-PCR).
Conclusions/Significance
The obtained goose transcriptome and DGE profiling data provide comprehensive gene expression information at the transcriptional level that could promote better understanding of the molecular mechanisms underlying follicle development and productivity.
doi:10.1371/journal.pone.0055496
PMCID: PMC3566205  PMID: 23405160
5.  BGI-RIS: an integrated information resource and comparative analysis workbench for rice genomics 
Nucleic Acids Research  2004;32(Database issue):D377-D382.
Rice is a major food staple for the world’s population and serves as a model species in cereal genome research. The Beijing Genomics Institute (BGI) has long been devoting itself to sequencing, information analysis and biological research of the rice and other crop genomes. In order to facilitate the application of the rice genomic information and to provide a foundation for functional and evolutionary studies of other important cereal crops, we implemented our Rice Information System (BGI-RIS), the most up-to-date integrated information resource as well as a workbench for comparative genomic analysis. In addition to comprehensive data from Oryza sativa L. ssp. indica sequenced by BGI, BGI-RIS also hosts carefully curated genome information from Oryza sativa L. ssp. japonica and EST sequences available from other cereal crops. In this resource, sequence contigs of indica (93-11) have been further assembled into Mbp-sized scaffolds and anchored onto the rice chromosomes referenced to physical/genetic markers, cDNAs and BAC-end sequences. We have annotated the rice genomes for gene content, repetitive elements, gene duplications (tandem and segmental) and single nucleotide polymorphisms between rice subspecies. Designed as a basic platform, BGI-RIS presents the sequenced genomes and related information in systematic and graphical ways for the convenience of in-depth comparative studies (http://rise.genomics.org.cn/).
doi:10.1093/nar/gkh085
PMCID: PMC308819  PMID: 14681438
6.  The Genomes of Oryza sativa: A History of Duplications 
Yu, Jun | Wang, Jun | Lin, Wei | Li, Songgang | Li, Heng | Zhou, Jun | Ni, Peixiang | Dong, Wei | Hu, Songnian | Zeng, Changqing | Zhang, Jianguo | Zhang, Yong | Li, Ruiqiang | Xu, Zuyuan | Li, Shengting | Li, Xianran | Zheng, Hongkun | Cong, Lijuan | Lin, Liang | Yin, Jianning | Geng, Jianing | Li, Guangyuan | Shi, Jianping | Liu, Juan | Lv, Hong | Li, Jun | Wang, Jing | Deng, Yajun | Ran, Longhua | Shi, Xiaoli | Wang, Xiyin | Wu, Qingfa | Li, Changfeng | Ren, Xiaoyu | Wang, Jingqiang | Wang, Xiaoling | Li, Dawei | Liu, Dongyuan | Zhang, Xiaowei | Ji, Zhendong | Zhao, Wenming | Sun, Yongqiao | Zhang, Zhenpeng | Bao, Jingyue | Han, Yujun | Dong, Lingli | Ji, Jia | Chen, Peng | Wu, Shuming | Liu, Jinsong | Xiao, Ying | Bu, Dongbo | Tan, Jianlong | Yang, Li | Ye, Chen | Zhang, Jingfen | Xu, Jingyi | Zhou, Yan | Yu, Yingpu | Zhang, Bing | Zhuang, Shulin | Wei, Haibin | Liu, Bin | Lei, Meng | Yu, Hong | Li, Yuanzhe | Xu, Hao | Wei, Shulin | He, Ximiao | Fang, Lijun | Zhang, Zengjin | Zhang, Yunze | Huang, Xiangang | Su, Zhixi | Tong, Wei | Li, Jinhong | Tong, Zongzhong | Li, Shuangli | Ye, Jia | Wang, Lishun | Fang, Lin | Lei, Tingting | Chen, Chen | Chen, Huan | Xu, Zhao | Li, Haihong | Huang, Haiyan | Zhang, Feng | Xu, Huayong | Li, Na | Zhao, Caifeng | Li, Shuting | Dong, Lijun | Huang, Yanqing | Li, Long | Xi, Yan | Qi, Qiuhui | Li, Wenjie | Zhang, Bo | Hu, Wei | Zhang, Yanling | Tian, Xiangjun | Jiao, Yongzhi | Liang, Xiaohu | Jin, Jiao | Gao, Lei | Zheng, Weimou | Hao, Bailin | Liu, Siqi | Wang, Wen | Yuan, Longping | Cao, Mengliang | McDermott, Jason | Samudrala, Ram | Wang, Jian | Wong, Gane Ka-Shu | Yang, Huanming
PLoS Biology  2005;3(2):e38.
We report improved whole-genome shotgun sequences for the genomes of indica and japonica rice, both with multimegabase contiguity, or almost 1,000-fold improvement over the drafts of 2002. Tested against a nonredundant collection of 19,079 full-length cDNAs, 97.7% of the genes are aligned, without fragmentation, to the mapped super-scaffolds of one or the other genome. We introduce a gene identification procedure for plants that does not rely on similarity to known genes to remove erroneous predictions resulting from transposable elements. Using the available EST data to adjust for residual errors in the predictions, the estimated gene count is at least 38,000–40,000. Only 2%–3% of the genes are unique to any one subspecies, comparable to the amount of sequence that might still be missing. Despite this lack of variation in gene content, there is enormous variation in the intergenic regions. At least a quarter of the two sequences could not be aligned, and where they could be aligned, single nucleotide polymorphism (SNP) rates varied from as little as 3.0 SNP/kb in the coding regions to 27.6 SNP/kb in the transposable elements. A more inclusive new approach for analyzing duplication history is introduced here. It reveals an ancient whole-genome duplication, a recent segmental duplication on Chromosomes 11 and 12, and massive ongoing individual gene duplications. We find 18 distinct pairs of duplicated segments that cover 65.7% of the genome; 17 of these pairs date back to a common time before the divergence of the grasses. More important, ongoing individual gene duplications provide a never-ending source of raw material for gene genesis and are major contributors to the differences between members of the grass family.
Comparative genome sequencing of indica and japonica rice reveals that duplication of genes and genomic regions has played a major part in the evolution of grass genomes
doi:10.1371/journal.pbio.0030038
PMCID: PMC546038  PMID: 15685292

Results 1-6 (6)