牦牛CSRP3基因的克隆及组织表达分析

Cloning and Tissue Expression Analysis of CSRP3 in Yak

CSRP3基因(Cysteine and glycine-rich protein 3,CSRP3)编码CRP3蛋白,是一个肌发生的正调节因子,可通过多种方式在肌肉发育和肌肉细胞结构维持中起重要作用。通过对牦牛CSRP3基因进行克隆及组织表达谱分析,为后续提高牦牛肉品质的研究提供基础数据。采用RT-PCR方法克隆牦牛CSRP3基因CDS区;再对其进行序列分析及蛋白结构和功能预测等生物信息学分析;最后利用实时荧光定量PCR技术检测该基因在牦牛不同组织中的表达量。牦牛CSRP3基因CDS区长585 bp,编码194个氨基酸;CSRP3基因的系统进化树结果显示,牦牛与黄牛的亲缘性最近,其次是绵羊。牦牛CSRP3基因编码的蛋白为偏碱性不稳定亲水蛋白,无跨膜结构和信号肽,为非分泌蛋白,含有磷酸化位点22个,N-糖基化位点2个,O-糖基化位点7个;存在两个LIM结构域,属于LIM结构域蛋白质超家族成员,主要分布于细胞核中;二级结构以无规卷曲为主,三级结构的最佳模型为1b8t.1.A;实时荧光定量PCR结果显示,牦牛CSRP3基因在臀大肌中有较高表达量。生物信息学分析结果显示,CRP3蛋白含有两个LIM结构域,主要分布在细胞核中,实时荧光定量PCR结果显示牦牛CSRP3基因在臀大肌中具有较高表达量,为牦牛CSRP3基因在牦牛肉品质方面的调控机制研究提供了基础数据。

The CSRP3 gene(cysteine and glycine-rich protein 3)encodes CRP3 protein,which is a positive regulator of myogenesis and plays an important role in muscle development and structural maintenance of muscle cells.Through cloning and tissue expression profile analysis of yak CSRP3 gene,the basic data may be provided for the further study of improving yak meat quality.The CDS region of yak CSRP3 gene was cloned by RT-PCR and analyzed by bioinformatics of sequence analysis,protein structure and function prediction.Finally,the expression of the gene in yak tissues was detected by qPCR.The results showed that the CDS region length of CSRP3 gene in yak was 585 bp,and a total of 194 amino acids were encoded.The phylogenetic tree demonstrated that yak had the closest relationship with cattle,followed by sheep.The protein encoded by CSRP3 gene was an alkaline unstable hydrophilic protein without transmembrane structure and signal peptide,not a secretory protein.There were 22 phosphorylation sites,2 N-glycosylation sites,7 O-glycosylation sites and 2 LIM domains,and it belonged to the superfamily member of LIM structure domain protein.The protein was mainly distributed in the nucleus.The secondary structure was mainly composed of random coils.The best model of the tertiary structure was 1b8t.1.A.The bioinformatics analysis revealed that the CRP3 protein contained 2 LIM domains and mainly distributed in the nucleus.The qPCR results showed that the yak CSRP3 gene had higher expression in the gluteus maximus.These results provide basic data for the research on whether the CSRP3 gene may improve the quality of yak meat.