牦牛<i>HDAC2</i>基因克隆及其在睾丸中的表达

doi:10.7668/hbnxb.20191525

华北农学报 ›› 2021, Vol. 36 ›› Issue (3): 222-229. doi: 10.7668/hbnxb.20191525

所属专题： 畜牧；生物技术；热点文章；

• 畜牧·水产·兽医 • 上一篇下一篇

牦牛HDAC2基因克隆及其在睾丸中的表达

殷实^1,2,3, 王斌¹, 曲尼拉姆¹, 杨柳青¹, 袁钰洁¹, 李键^1,2,3

1. 西南民族大学青藏高原动物遗传资源保护与利用教育部重点实验室, 四川成都 610041;
2. 西南民族大学青藏高原动物遗传资源保护与利用四川省重点实验室, 四川成都 610041;
3. 西南民族大学畜牧兽医学院, 四川成都 610041;
4. 西南民族大学现代生物技术国家民委重点实验室, 四川成都 610041

收稿日期:2021-03-04 出版日期:2021-06-28
通讯作者: 李键(1967-)，男，四川理县人，教授，博士，主要从事牦牛细胞生物学和发育生物学研究。
作者简介:殷实(1988-)，男，河南焦作人，讲师，博士，主要从事动物细胞与胚胎工程研究。
基金资助:
中央高校基本科研业务费专项基金项目（2021NYB01）

Cloning of Yak HDAC2 Gene and Its Expression Pattern in Testis

YIN Shi^1,2,3, WANG Bin¹, QUNI Lamu¹, YANG Liuqing¹, YUAN Yujie¹, LI Jian^1,2,3

1. Key Laboratory of Qinghai-Tibetan Plateau Animal Genetic Resource Reservation and Utilization of Ministry of Education, Southwest Minzu University, Chengdu 610041, China;
2. Qinghai-Tibetan Plateau Animal Genetic Resource Reservation and Utilization Key Laboratory of Sichuan Province, Southwest Minzu University, Chengdu 610041, China,;
3. College of Animal & Veterinary Sciences, Southwest Minzu University, Chengdu 610041, China;
4. Key Laboratory of Modern Biotechnology of State Ethnic Affairs Commission, Southwest Minzu University, Chengdu 610041, China

Received:2021-03-04 Published:2021-06-28

摘要/Abstract

摘要： 旨在克隆牦牛HDAC2（Histone deacetylase 2）基因，预测分析HDAC2蛋白的结构和特性，并检测HDAC2的mRNA在牦牛不同组织以及不同发育阶段睾丸中的表达。将牦牛按年龄划分为幼年组（0.5～1岁）、成年组（4～5岁）及老年组（7～9岁），采集成年组牦牛肝脏、肾脏、肺、胃、脾脏、脑、心脏和卵巢组织以及3个时期的牦牛睾丸组织，利用反转录PCR （RT-PCR）技术克隆获得牦牛HDAC2的基因序列，使用生物信息学的方法预测该基因的序列同源性以及其编码蛋白质的氨基酸序列和结构；通过实时荧光定量PCR检测HDAC2的mRNA在牦牛不同组织中的表达，通过RT-PCR检测HDAC2的mRNA在牦牛不同发育阶段睾丸中的表达，利用色素原位杂交技术检测HDAC2 mRNA在成年牦牛睾丸中的定位。结果表明，HDAC2基因的开放阅读框包含1 467个碱基，编码488个氨基酸，其基因序列在哺乳动物中高度保守。结构预测表明，HDAC2是一个脂溶性疏水蛋白，具有一个组蛋白去乙酰化酶结构域。该基因的mRNA在卵巢和睾丸中的表达较高。HDAC2的mRNA在幼年期牦牛睾丸中的表达显著高于其他时期，在牦牛睾丸中HDAC2的mRNA定位在除精细胞之外的各类细胞中。克隆得到牦牛HDAC2基因序列并明确了该基因在睾丸中的时序表达模式，对于深入探讨HDAC2在牦牛睾丸发育及精子发生中发挥的作用提供了一定的试验数据。

关键词: 牦牛, 组蛋白去乙酰化, HDAC2, 基因克隆, 睾丸

Abstract: The aim of this research is to clone the HDAC2 gene in yak, predict the structure and property of HDAC2 protein, and identify the expression profile of HDAC2 mRNA in different tissues and testis of different stages of yak. Yaks were divided into juvenile(0.5-1 year), adult(4-5 years) and old groups(7-9 years).The samples of liver, kidney, lung, stomach, spleen, brain, heart and ovary from adult group and testis of three stages were collected. The yak HDAC2 gene was cloned by reverse transcription PCR (RT-PCR). The homology of HDAC2 gene sequence, as well as the amino acid sequence and structure of HDAC2 protein were predicated by bioinformatics. Real-time PCR was applied to identify the mRNA expression profile of HDAC2 in different tissues of yak. RT-PCR was applied to detect the mRNA expression of HDAC2 in yak testis of different stages. Chromogenic in situ hybridization (CISH) was applied to analysis the localization of HDAC2 mRNA in adult yak testis. Result showed that the open reading frame (ORF) of yak HDAC2 contained 1 467 bases and encoded 488 amino acids. The sequence of yak HDAC2 was highly conserved in mammals. Structure prediction indicated that HDAC2 was a hydrophilic liposoluble protein with a histone deacetylase domain. The mRNA of HDAC2 was highly expressed in ovary and testis. The expression of HDAC2 in juvenile stage was significantly higher than that in other stages. The mRNA of HDAC2 localized in multiple kinds of testicular cells except spermatid. The sequence of yak HDAC2 gene was cloned and the spatiotemporal expression pattern of this gene in testis was verified. This study provides some experimental data for further study on the role of HDAC2 in testicular development and spermatogenesis of yak.

Key words: Yak, Histone deacetylation, HDAC2, Gene cloning, Testis

中图分类号:

Q78
S823

殷实, 王斌, 曲尼拉姆, 杨柳青, 袁钰洁, 李键. 牦牛HDAC2基因克隆及其在睾丸中的表达[J]. 华北农学报, 2021, 36(3): 222-229. doi: 10.7668/hbnxb.20191525.

YIN Shi, WANG Bin, QUNI Lamu, YANG Liuqing, YUAN Yujie, LI Jian. Cloning of Yak HDAC2 Gene and Its Expression Pattern in Testis[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2021, 36(3): 222-229. doi: 10.7668/hbnxb.20191525.

http://www.hbnxb.net/CN/Y2021/V36/I3/222

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