本文選題：HSP基因 + 天祝白牦牛��； 參考：《核農(nóng)學(xué)報(bào)》2017年07期

【摘要】：熱休克蛋白60(HSP60)不僅在細(xì)胞保護(hù)方面發(fā)揮作用,也參與雄性動(dòng)物生殖生理的調(diào)控。為研究HSP60基因序列特性及其在天祝白牦牛下丘腦-垂體-睪丸軸(HPT-axis)上的表達(dá)情況,探尋其在白牦牛睪丸發(fā)育、精子發(fā)生過(guò)程中的作用機(jī)制,本研究采用RT-PCR和RACE技術(shù)獲得白牦牛HSP60基因全長(zhǎng)cDNA序列,采用生物信息學(xué)方法分析HSP60蛋白的理化性質(zhì)、結(jié)構(gòu)及不同物種之間的同源性等,并利用qPCR、Western blotting、免疫組化法分析HSP60在白牦牛下丘腦-垂體-睪丸軸中的表達(dá)及定位。結(jié)果表明,白牦牛HSP60基因cDNA全長(zhǎng)為2 300 bp,開放閱讀框?yàn)? 722 bp,編碼572個(gè)氨基酸;其理論分子量為60.977k Da、等電點(diǎn)為5.69,編碼的蛋白為非跨膜可溶性蛋白。氨基酸序列比對(duì)結(jié)果顯示,白牦牛HSP60氨基酸序列與牛、瘤牛、綿羊、藏羚羊、駱駝、白犀牛、兔和黑猩猩的氨基酸序列同源性高、進(jìn)化水平相近。HSP60基因及蛋白在白牦牛的下丘腦、垂體及睪丸組織中均有表達(dá),其中下丘腦及垂體組織表達(dá)量顯著高于睪丸,下丘腦和垂體之間差異不顯著。免疫組化結(jié)果顯示,HSP60蛋白定位表達(dá)于白牦牛下丘腦組織的室旁核大細(xì)胞、室旁核小細(xì)胞和神經(jīng)角演網(wǎng)、垂體組織的腺細(xì)胞、睪丸組織的精原細(xì)胞、精母細(xì)胞、支持細(xì)胞和間質(zhì)細(xì)胞,而精子細(xì)胞中表達(dá)較弱。通過(guò)對(duì)健康成年白牦牛HSP60在下丘腦-垂體-睪丸軸的表達(dá)與定位檢測(cè),推斷HSP60參與雄性白牦牛生殖軸調(diào)控,并參與睪丸發(fā)育與精子發(fā)生。本研究結(jié)果為進(jìn)一步研究HSP60對(duì)雄性生殖調(diào)控奠定了基礎(chǔ)。
[Abstract]:Heat shock protein 60 (HSP60) not only plays a role in cell protection, but also participates in the regulation of reproductive physiology in male animals. To study the sequence characteristics of HSP60 gene and its expression in the hypothalamus-pituitary-testicular axis of Tianzhu white yak, and to explore the role of HSP60 gene in the development of testis and spermatogenesis of white yak. In this study, the full-length cDNA sequence of HSP60 gene of white yak was obtained by RT-PCR and race. The physicochemical properties, structure and homology of HSP60 protein were analyzed by bioinformatics. The expression and localization of HSP60 in the hypothalamus-pituitary-testicular axis of white yak were analyzed by using qPCRX Western blotting and immunohistochemistry. The results showed that the full length of HSP60 gene cDNA of white yak was 2 300 BP, the open reading frame was 1 722 BP, the theoretical molecular weight was 60.977 K Da, the isoelectric point was 5.69, and the protein was non-transmembrane soluble protein. The results of amino acid sequence alignment showed that the amino acid sequence of HSP60 in white yak had high homology with cattle, sheep, Tibetan antelope, camel, white rhinoceros, rabbit and chimpanzee. HSP60 gene and protein were expressed in hypothalamus, pituitary and testis of white yak, and the expression of hypothalamus and pituitary was significantly higher than that of testis, but there was no significant difference between hypothalamus and pituitary. Immunohistochemical results showed that HSP60 protein was localized in the large paraventricular nucleus cells, paraventricular nucleus small cells and neural keratinocytes, pituitary gland cells, testicular spermatogonia and spermatocytes in the hypothalamus of white yak. Sertoli cells and interstitial cells were less expressed in sperm cells. By detecting the expression and localization of HSP60 in the hypothalamus-pituitary-testicular axis of healthy adult white yaks, it was inferred that HSP60 was involved in the regulation of reproductive axis, testicular development and spermatogenesis of male white yaks. The results laid a foundation for further study on the regulation of HSP60 on male reproduction.
【作者單位】： 甘肅農(nóng)業(yè)大學(xué)動(dòng)物醫(yī)學(xué)院;甘肅農(nóng)業(yè)大學(xué)動(dòng)物科技學(xué)院;
【基金】：“十二五”農(nóng)村領(lǐng)域國(guó)家科技計(jì)劃項(xiàng)目(2013AA02505-3)
【分類號(hào)】：Q78;S823.85

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本文編號(hào)：2028969