牦牛Myf6基因克隆及生物信息学分析

doi:10.16431/j.cnki.1671-7236.2019.06.005

摘要/Abstract

摘要：

本试验旨在克隆牦牛Myf6基因，进行生物信息学分析并探究其在牦牛不同时期的表达情况。以青海雪多牦牛为研究对象，通过设计引物对其Myf6基因CDS区进行克隆测序，运用生物信息学软件对其功能结构进行预测研究。结果显示，牦牛Myf6基因含有一个大小为729 bp的开放阅读框（ORF），编码242个氨基酸。同源性比对和进化树分析结果表明，牦牛Myf6氨基酸序列与普通牛、绵羊亲缘性最近。对Myf6蛋白理化性质进行预测分析得到其结构式为C₁₁₆₇H₁₈₇₀N₃₃₆O₃₇₂S₁₃，理论等电点为5.64，其中丝氨酸含量最高，其次是亮氨酸、谷氨酸、脯氨酸和精氨酸；其疏水平均值为-0.586，亲水性最强为-2.467，疏水性最强为1.511，为亲水性、可溶性蛋白；有可形成卷曲螺旋部位；磷酸化位点预测其有26个丝氨酸激酶，10个苏氨酸激酶，5个酪氨酸激酶；亚细胞定位分析其编码产物在细胞核中分布最多（73.9%），其次为细胞骨架（13.0%）、细胞质（4.3%）、高尔基体（4.3%）、分泌系统小泡（4.3%）。蛋白质二级结构主要由无规则卷曲（49.17%）组成，其次是α-螺旋（33.47%）、延伸连（11.57%）及β-折叠（5.79%）。蛋白功能分析显示其生长因子功能几率较大（7.694），其次为转录调节作用，预测其作为一种生长因子参与机体生物学调控。实时荧光定量PCR结果显示，Myf6基因在胎牛、6月龄、4~5岁牦牛背最长肌中均有表达，且在6月龄的表达量显著高于胎牛和4~5岁牦牛（P<0.05）。上述结果为研究牦牛Myf6生肌因子提供了重要的研究数据，为改良牦牛经济性状提供了参考。

关键词: 牦牛; Myf6基因; 生物信息学分析; 克隆

Abstract:

The aim of this study was to clone and analyze Myf6 gene in yaks by bioinformatics,and investigate its expression in different periods.The snowy yak in Qinghai was taken as the research object.The Myf6 gene CDS region was cloned and sequenced by designing primers,and its functional structure was predicted by bioinformatics software.The results showed that the yak Myf6 gene contained a 729 bp open reading frame (ORF) encoding 242 amino acids.The results of homology alignment and phylogenetic tree analysis indicated that the amino acid sequence of yak Myf6 was close to common cattle and sheep.The physicochemical properties of Myf6 protein were predicted and analyzed.The structural formula of Myf6 protein was C₁₁₆₇H₁₈₇₀N₃₃₆O₃₇₂S₁₃,and the theoretical isoelectric point was 5.64,with serine being the highest,followed by leucine,glutamic acid,proline and arginine.The hydrophobic average was -0.586,the most hydrophilic was -2.467,and the hydrophobicity was 1.511.It was predicted that the yak Myf6 was predicted to be hydrophilic and soluble protein.There were portions that could form coiled coil;The phosphorylation sites results showed that there were 26 serine kinases,10 threonine kinases and 5 tyrosine kinases;Subcellular localization analysis showed that the encoded product was most distributed in the nucleus (73.9%),followed by cytoskeleton (13.0%),cytoplasm (4.3%),Golgi apparatus (4.3%) and secretory system vesicles (4.3%).The protein secondary structure was predicted to consist mainly of random coils (49.17%),followed by alpha helix (33.47%),extended chain (11.57%) and beta-sheet (5.79%).Functional analysis of the protein showed that the growth factor function had a higher probability (7.694),followed by transcriptional regulation,which was predicted to participate in the biological regulation of the body as a growth factor.The results of Real-time PCR showed that Myf6 gene was expressed in the longissimus dorsi muscle of fetal calf,6 months old and 4 to 5 years old,and the expression level at 6 months old was significantly higher than that of fetal cattle and 4 to 5 years old (P<0.05).The above results provided important research data for the study of yak Myf6 myogenic factors,and provided a reference for improving the economic traits of yak.

Key words: yak; Myf6 gene; bioinformatics analysis; cloning

中图分类号:

S823.8⁺5

雷秦祎, 付东海, 常永芳, 吴晓云, 梁春年, 褚敏, 阎萍. 牦牛Myf6基因克隆及生物信息学分析[J]. 《中国畜牧兽医》, 2019, 46(6): 1603-1611.

LEI Qinyi, FU Donghai, CHANG Yongfang, WU Xiaoyun, LIANG Chunnian, CHU Min, YAN Ping. Cloning and Bioinformatics Analysis of Myf6 Gene in Yak[J]. , 2019, 46(6): 1603-1611.

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