秦川牛CAP2基因CDS区克隆及表达谱分析

doi:10.12160/j.issn.1672-5190.2022.03.003

摘要/Abstract

摘要： [目的]克隆秦川牛CAP2基因的编码区序列（CDS）,分析该基因在不同组织及原代脂肪细胞分化过程中的表达特征。[方法]采用RT-PCR方法扩增秦川牛CAP2基因的CDS区,运用ProtPram、TMpred、ProtFun 2.1 Server等在线网站进行CAP2蛋白的生物信息学分析;通过油红O染色和qPCR检测成脂标志基因PPARγ和FABP4基因的表达水平以构建牛原代脂肪细胞诱导分化体系;利用qPCR检测分析CAP2基因在秦川牛7种组织（心、肝、脾、肺、肾、肌肉和背脂）和原代脂肪细胞分化过程（0~10 d）中的表达。[结果]秦川牛CAP2基因CDS区长1 461 bp,编码486 个氨基酸,氨基酸序列主要由无规卷曲和α-螺旋构成。CAP2基因在脂肪组织中高表达,极显著（P<0.01）高于其他组织。牛原代脂肪细胞诱导分化过程中,PPARγ和FABP4基因的表达量逐渐上升,与第0天相比第10 天时表达量达到最大（P<0.01）;与对照组相比,诱导分化组脂滴积累明显增加;CAP2基因表达量也随时间推移逐渐上升,以0 d为对照,第10天表达量最高（P<0.001）。[结论]成功克隆了秦川牛CAP2基因全长1 461 bp的编码区;CAP2基因在牛脂肪组织中以较高水平表达,且CAP2基因可能参与脂肪生成与分化过程,预测CAP2基因可能是促进成脂分化的转录因子,对维持牛脂肪细胞状态发挥关键作用,可能作为秦川牛肉质性状的候选基因,该研究结果为进一步揭示牛CAP2基因的功能提供基础资料。

关键词: 秦川牛, CAP2基因, 脂肪细胞, 表达模式, 生物信息学分析

Abstract: [Objective] To clone the coding region sequence （CDS） of CAP2 gene in Qinchuan bovine, and to characterize the expression profiling of this gene in various tissues and during the primary adipocyte differentiation. [Method] The CDS region of CAP2 gene in Qinchuan bovine was amplified by RT-PCR assay, and the CAP2 protein was analyzed using online bioinformatical tools such as ProtPram, TMpred, and ProtFun 2.1 Server. The gene expression levels of PPARγ and FABP4 were detected using qPCR assay and oil red O staining method to construct a bovine primary adipocyte induced differentiation system. Moreover, the expression levels of CAP2 gene in seven tissues （heart, liver, spleen, lung, kidney, muscle, and back fat） as well as during the differentiation process of primary adipocyte （0-10 days） in Qinchuan bovine were assessed using qPCR assay. [Result] The CDS region of CAP2 gene in Qinchuan bovine was 1 461 bp and encoded 486 amino acids. Its amino acid sequence mainly composed of random coil and α-helix. Extremely significantly （P<0.01） higher expression level of CAP2 gene was observed in back fat in comparison to the other tested tissues. During the differentiation process of primary adipocyte, the expression levels of PPARγ and FABP4 genes increased gradually and peaked on day 10, which were extremely significantly （P<0.01） higher compared to day 0. The induced differentiation group of primary adipocyte had a much larger lipid droplet accumulation than the control group. Furthermore, the expression level of CAP2 gene increased gradually with the prolongation of induced differentiation duration, and peaked on day 10 as well, which were extremely significantly （P<0.001） higher compared to day 0. [Conclusion] The 1 461 bp full-length CDS of CAP2 gene in Qinchuan bovine has been successfully cloned. The CAP2 gene is highly expressed in adipose tissue and may involves in adipogenesis and differentiation. As a result, the CAP2 gene is assumed to be a transcription factor that promotes adipogenic differentiation, plays a key role in regulating the status of bovine adipocyte, and could be a candidate gene associated with meat quality trait of Qinchuan bovine. These findings provide preliminary information for further revealing the function of bovine CAP2 gene.

Key words: Qinchuan bovine, CAP2 gene, adipocyte, expression pattern, bioinformatic analysis

中图分类号:

S823.2

高玉红, 户春丽, 马燕芬, 汪书哲, 马云. 秦川牛CAP2基因CDS区克隆及表达谱分析[J]. 畜牧与饲料科学, 2022, 43(3): 16-24.

GAO Yu-hong, HU Chun-li, MA Yan-fen, WANG Shu-zhe, MA Yun. Cloning and Expression Profiling of CDS Region of CAP2 Gene in Qinchuan Bovine[J]. Animal Husbandry and Feed Science, 2022, 43(3): 16-24.

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