本文選題：蒙古馬　切入點(diǎn)：睪丸　出處：《內(nèi)蒙古農(nóng)業(yè)大學(xué)》2017年碩士論文

【摘要】：蒙古馬是世界上最古老的馬品種之一。具有良好的繁殖能力和獨(dú)特的遺傳基因。通過測量馬睪丸可有效預(yù)測種公馬的繁殖能力。睪丸發(fā)育是雄性哺乳動(dòng)物重要的生理過程。產(chǎn)生精子和分泌雄性激素是睪丸的基本功能。為了從基因水平了解性成熟前后蒙古馬睪丸發(fā)育及精子生成的調(diào)控機(jī)制,本實(shí)驗(yàn)采用新一代高通量測序技術(shù),對(duì)蒙古馬性成熟前后兩個(gè)發(fā)育階段的睪丸組織建立轉(zhuǎn)錄組文庫進(jìn)行測序、對(duì)結(jié)果進(jìn)行可變剪切分析及差異表達(dá)基因分析;對(duì)差異表達(dá)基因進(jìn)行GO功能富集分析、KEGG Pathway分析、轉(zhuǎn)錄因子分析,找到睪丸發(fā)育及精子生成相關(guān)GO功能條目、KEGG通路及相關(guān)基因。最后應(yīng)用實(shí)時(shí)熒光定量PCR方法檢測蒙古馬睪丸中TSSK2、SPATA6、CDC20、MAD2L1基因的mRNA表達(dá)量在性成熟前和性成熟后的變化。本研究對(duì)于選擇優(yōu)良種用公馬具有重要意義。揭示公馬睪丸發(fā)育及精子生成的分子機(jī)制,并獲得以下研究結(jié)果:(1)經(jīng)過對(duì)蒙古馬性成熟前后期進(jìn)行轉(zhuǎn)錄組分析,得到24878個(gè)差異表達(dá)基因,其中上調(diào)基因16237個(gè),下調(diào)基因8641個(gè)。這些差異基因被注釋到10019條GO功能條目上。主要富集在精子形成(spermatogenesis)、雄性配子的一代(male gamete generation)、精細(xì)胞的發(fā)育(spermatid development)等與睪丸發(fā)育及精子生成緊密相關(guān)的生物學(xué)過程。(2)對(duì)差異表達(dá)基因進(jìn)行KEGG通路分析發(fā)現(xiàn):與睪丸發(fā)育及精子生成緊密相關(guān)的通路有卵母細(xì)胞減數(shù)分裂(Oocyte meiosis)、黃體酮調(diào)節(jié)的卵母細(xì)胞減數(shù)分裂(Progesterone-mediated oocyte maturation)等通路。(3)采用實(shí)時(shí)熒光定量PCR技術(shù)對(duì)TSSK2基因、SPATA6基因、CDC20基因及MAD2L1基因進(jìn)行驗(yàn)證,結(jié)果顯示以上4個(gè)基因在性成熟后睪丸中的表達(dá)量比性成熟前睪丸中的表達(dá)量高2倍以上,且差異顯著。證明了轉(zhuǎn)錄組測序結(jié)果的準(zhǔn)確性。
[Abstract]:Mongolian horse is one of the oldest horse breeds in the world.Has good reproduction ability and unique genetic genes.By measuring the testis of the horse, the breeding ability of the male horse can be predicted effectively.Testicular development is an important physiological process in male mammals.The production of sperm and the secretion of testosterone are the basic functions of the testis.In order to understand the regulatory mechanism of testis development and spermatogenesis in Mongolian horses before and after sexual maturation at the gene level, a new generation of high-throughput sequencing technique was used.The transcriptional library of testis in two stages of sexual maturation was sequenced, the results were analyzed by variable shear analysis and differential expression gene analysis, and the differentially expressed genes were analyzed by go functional enrichment analysis and KEGG Pathway analysis.Transcription factor analysis revealed KEGG pathway and related genes related to testicular development and spermatogenesis.The changes of mRNA expression of TSSK2SPATA6SPATA6CDC20 / MAD2L1 gene in the testis of Mongolian horses before and after sexual maturation were detected by real-time fluorescence quantitative PCR.This study is of great significance for the selection of a good breed of male horse.The molecular mechanism of testicular development and spermatogenesis in male horses was revealed, and the following results were obtained: 1) 24878 differentially expressed genes were obtained by transcriptome analysis before and after sexual maturation of Mongolian horses, including 16237 up-regulated genes and 8641 down-regulated genes.These differential genes were annotated to 10019 go functional entries.The differential expression genes were mainly enriched in spermatogenesis, male gamete generation gamete generation, spermatocyte development and spermatid development and other biological processes closely related to testicular development and spermatogenesis. The results of KEGG pathway analysis of differentially expressed genes were as follows:The pathways closely related to development and spermatogenesis include oocyte meiosism, progesterone-mediated oocyte maturation.3.) the TSSK2 gene, SPATA6 gene, CDC20 gene and MAD2L1 gene were verified by real-time fluorescence quantitative PCR.The results showed that the expression of the above four genes in the testis after sexual maturation was more than 2 times higher than that in the testis before sexual maturation, and the difference was significant.The results of transcriptome sequencing were proved to be accurate.
【學(xué)位授予單位】：內(nèi)蒙古農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：S821.81

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