摘要
【目的】探明牦牛和犏牛的附睾尾部差异表达基因(DEGs),筛选出与精子成熟和贮存密切相关的功能基因,为揭示犏牛精子发生及其成熟过程的分子机制打下理论基础。【方法】以牦牛和犏牛的附睾尾部为研究对象,通过Illumina 127-HiSeq 2000平台完成高通量转录组测序,经过滤、质量控制及拼接组装后,依据FDR<0.05且|log_(2)Fold Change|>1的筛选标准,通过EBSeq筛选出DEGs,然后进行GO功能注释分析和KEGG信号通路富集分析,并以实时荧光定量PCR验证高通量转录组测序数据的准确性。【结果】在牦牛和犏牛的附睾尾部共筛选获得76个DEGs,其中43个DEGs上调、33个DEGs下调;76个DEGs在COG、GO、KEGG、KOG、Nr、Pfam、Swiss-Prot和eggNOG等数据库中均有注释信息,尤其在COG和Pfam数据库中的注释率最高(达88.16%)。GO功能注释分析结果显示,DEGs被注释到生物过程(Biological process)、细胞组分(Cellular component)和分子功能(Molecular function)三大功能类别上;KEGG信号通路富集分析发现76个DEGs主要显著富集在3条信号通路上,分别是胆汁分泌通路(ko04976:Bile secretion)、ABC转运器(ko02010:ABC transporters)和cAMP信号通路(ko04024:cAMP signaling pathway)。随机选择8个DEGs(SERPINA1、MMP7、ATP2C1、ABCC1、NMT1、NAT1、CFTR和PRX)进行实时荧光定量PCR检测验证,结果显示这8个DEGs的表达模式与高通量转录组测序的结果基本一致,表明转录组数据准确可靠。【结论】在牦牛和犏牛的附睾尾部存在76个DEGs(43个DEGs上调,33个DEGs下调),显著富集在胆汁分泌通路、ABC转运器及c AMP信号通路上,与精子获能相关的DEGs有MMP7、IGFBP2和ABCC4基因,且这3个基因在犏牛附睾尾部呈下调表达,即精子获能失败可能是导致犏牛雄性不育的主要原因。
【Objective】In order to find out the differentially expressed genes(DEGs)in the epididymal cauda of yak and cattleyak,functional genes closely related to sperm maturation and storage were excavated,which laid a theoretical foundation for exploring the molecular mechanism of spermatogenesis and maturation process in cattleyak.【Method】Ta-king the epididymal cauda of yaks and cattleyaks as the research subject,high-throughput transcriptome sequencing was completed by Illumina 127-HiSeq 2000 platform,and after filtration,quality control and splicing assembly,DEGs were screened out by EBSeq according to the screening criteria of FDR<0.05 and|log_(2) Fold Change|>1,and then GO function annotation analysis and KEGG signal pathway enrichment analysis were performed.The accuracy of high-throughput tran-scriptome sequencing data was verified by real-time fluorescence quantitative PCR.【Result】A total of 76 DEGs were screened in the caput of the epididymis of yaks and calves,of which 43 DEGs were up-regulated and 33 DEGs were down-regulated.76 DEGs had annotation information in COG,GO,KEGG,KOG,NR,PFAM,Swiss-Prot and egg-NOG databases,especially the highest annotation rate in the COG and Pfam databases(88.16%).The results of GO func-tion annotation analysis showed that DEGs were annotated into three functional categories:biological process,cellular component and molecular function.KEGG signaling pathway enrichment analysis found that 76 DEGs were mainly signifi-cantly enriched in 3 signaling pathways,namely bile secretion pathway(ko04976:Bile secretion),ABC transporter(ko02010:ABC transporters)and cAMP signaling pathway(ko04024:cAMP signaling pathway).Eight DEGs(SERPI-NA1,MMP7,ATP2C1,ABCC1,NMT1,NAT1,CFTR,PRX)were randomly selected for real-time fluorescence quanti-tative PCR,and the expression patterns of the eight DEGs were basically consistent with the results of high-throughput transcriptome sequencing,indicating that the transcriptome data were accurate and reliable.【Conclusion】There are 76 DEGs(43 DEGs up-
作者
赵旺生
李柯锐
张婷婷
潘美兰
王鹏
李春海
张鹏
张永德
ZHAOWang-sheng;LI Ke-rui;ZHANG Ting-ting;PAN Mei-lan;WANG Peng;LI Chun-hai;ZHANG Peng;ZHANG Yong-de(School of Life Science and Engineering,Southwest University of Science and Technology,Mianyang,Sichuan 621000,China;Key Discipline Laboratory of National Defense for NuclearWaste and Environmental Security,Southwest University of Science and Technology,Mianyang,Sichuan 621000;Golog Snow Mountain Food Co.,Ltd.,Golog,Qinghai 814099,China)
出处
《南方农业学报》
CAS
CSCD
北大核心
2023年第5期1273-1282,共10页
Journal of Southern Agriculture
基金
国家自然科学基金青年基金项目(31601946)
青海省中央引导地方科技发展资金项目(2023ZY026)
四川省科技计划项目(2021YFH0101)。
关键词
牦牛
犏牛
附睾尾部
差异表达基因(DEGs)
精子
雄性不育
高通量转录组测序
yak
cattleyak
epididymal cauda
differentially expressed genes(DEGs)
sperm
male sterility
high-throughput transcriptome sequencing
