牦牛StAR基因克隆及其生物信息学与组织表达特性的研究

doi:10.11843/j.issn.0366-6964.2021.09.008

摘要/Abstract

摘要： 旨在克隆获得牦牛StAR基因编码序列（CDS）并进行生物信息学分析，探究其mRNA组织表达特性。本研究以屠宰场采集的成年母牦牛心、肝、脾、肺、肾、卵巢、输卵管、子宫组织（n=5），不同年龄（胎牛、1岁、2岁）牦牛的卵巢（n=3），不同发情周期（卵泡期、黄体期）的牦牛卵巢（n=3），黄体期黄牛的卵巢（n=3）及实验室冻存的牦牛颗粒细胞为研究材料。以牦牛黄体期卵巢cDNA为模板，用逆转录PCR克隆StAR基因，并使用MEGA7.0和ExPASy-ProtParam等软件分析其生物信息学特性；采用实时荧光定量PCR技术分析牦牛StAR基因组织表达特性。结果发现，StAR基因CDS区长858 bp，编码285个氨基酸，StAR蛋白总体带正电荷，属于碱性亲水稳定蛋白，无跨膜结构及信号肽，主要存在于细胞质和线粒体； StAR基因具有较高的保守性，符合物种进化规律。牦牛StAR基因在卵巢表达水平最高（P<0.01），且2岁时卵巢表达水平极显著高于胎牛和1岁龄（P<0.01），黄体期卵巢表达水平极显著高于卵泡期（P<0.01）；黄体期黄牛卵巢中StAR基因的表达量极显著高于牦牛（P<0.01）；在颗粒细胞的体外培养过程中StAR基因表达量逐渐上升，在培养24 h时达到高峰（P<0.01），随后显著降低。综上所述，StAR基因序列较为保守，在牦牛卵巢组织中表达最高，且表达水平随年龄与卵巢周期而变化，提示StAR基因可能参与牦牛卵巢及黄体功能相关的繁殖调控。

关键词: 牦牛, StAR基因, 基因克隆, 组织表达, 卵巢

Abstract: The objective of this study was to obtain the coding sequence (CDS) of yak StAR gene, conduct bioinformatics analysis and investigate its mRNA expression characteristics in different tissues. Yak samples were collected from local slaughter house, including heart, liver, spleen, lung, kidney, ovary, oviduct and uterus of adult female yaks (n=5), ovary of yaks at different ages (fetal, 1-year-old and 2-years-old) (n=3), ovary of yaks during different periods of estrous cycles (follicular phase and luteal phase) (n=3), the luteal phase ovary of cattle (n=3). Granulosa cells of yaks were cryopreserved previously. StAR gene was cloned using yak ovarian cDNA as template during luteal phase by reverse transcriptase-polymerase chain reaction (PCR) and its bioinformatics properties were analyzed by bioinformatics software such as MEGA7.0 and ExPASy-ProtParam etc. The tissue expression characteristics of yak StAR gene were detected by quantitative real-time PCR. The results showed that the CDS of yak StAR gene was 858 bp, encoding 285 amino acids. StAR protein was positively charged and belonged to alkaline hydrophilic stable protein, which had no transmembrane structure and signal peptide. It mainly existed in cytoplasm and mitochondria. StAR gene was highly conserved between different species, which accorded with the law of species evolution. The expression level of yak StAR gene was the highest in ovary (P<0.01). The expression level of StAR gene in the ovary of 2-years-old was significantly higher than that of fetal and 1-year-old (P<0.01). Its expression in yak ovary during luteal phase was significantly higher than that during follicular phase (P<0.01), and was significantly higher in cattle ovary than in yak ovary during luteal phase(P<0.01). During the in vitro culture (IVC) of granulosa cells, the expression of StAR gene increased gradually and reached the peak level at 24 h IVC (P<0.01), and then significantly decreased. The results showed that StAR gene was relatively conservative in the course of animal evolution. Its expression level was greatest in the ovary of yak, and changing with age- and estrous cycle-specific characteristics, which indicate that StAR gene may be involved in the regulation of yak reproduction associated with ovary and corpus luteum function.

Key words: yak, StAR gene, gene cloning, tissue expression, ovary

中图分类号:

S823.85

刘宇, 邬建飞, 李恒, 荆天, 卢建远, 字向东. 牦牛StAR基因克隆及其生物信息学与组织表达特性的研究[J]. 畜牧兽医学报, 2021, 52(9): 2452-2463.

LIU Yu, WU Jianfei, LI Heng, JING Tian, LU Jianyuan, ZI Xiangdong. Cloning and Analysis of Bioinformatics and Tissue Expression Characteristics of Yak StAR Gene[J]. Acta Veterinaria et Zootechnica Sinica, 2021, 52(9): 2452-2463.

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