摘要：

为探究牦牛Akirin1基因的序列特征,详细了解其组织表达特性。以甘南牦牛肌肉组织cDNA为模板,利用RT-PCR技术克隆甘南牦牛Akirin1基因CDS序列,并使用多种生物在线软件及工具进行生物信息学分析,再利用qPCR技术检测Akirin1基因在甘南牦牛心、肝、脾、肺、肾、肌肉、脂肪和睾丸组织中表达差异。结果显示,甘南牦牛Akirin1基因CDS长579 bp,共编码192个氨基酸,基因编码的蛋白分子式为C₉₆₄H₁₅₂₅N₂₇₅O₂₉₁S₈,蛋白理论等电点为8.91,不稳定系数和总平均亲水性分别为84.14,-0.822,属于不稳定亲水性蛋白;蛋白潜在磷酸化位点有29个,分别为14个丝氨酸,11个苏氨酸和4个酪氨酸;无信号肽和跨膜螺旋结构,亚细胞定位显示,主要存在于细胞核中,属于非分泌型蛋白;蛋白高级结构主要由39.06%的α-螺旋和56.25%的无规则卷曲结构组成。同源性分析比对牦牛与普通牛亲缘关系最近,符合实际情况,说明Akirin1基因在进化过程中相对具有一定保守性。实时荧光定量结果显示,Akirin1基因在成年甘南牦牛心、肝、脾、肺、肾、肌肉和脂肪组织均有所表达,而在肾脏组织中相对表达最多,显著高于其他组织,在肌肉和肝脏组织中表达相对较少。本试验为进一步研究牦牛Akirin1基因功能特征提供一定的理论基础。

关键词: 牦牛, Akirin1基因, 克隆, 生物信息学分析, 组织表达分析

Abstract:

In order to explore the sequence characteristics of yak Akirin1 gene and understand its tissue expression characteristics in detail.Gannan yak muscle tissue cDNA was used as a template to clone the CDS sequence of Gannan yak Akirin1 gene by RT-PCR technology,and bioinformatics analysis was performed using a variety of biological online software and tools.The qPCR technique was used to detect the difference expression of Akirin1 gene in Gannan yak heart,liver,spleen,lung,kidney,muscle,fat and testis tissues.The results showed that the CDS of Akirin1 gene in Gannan yak was 579 bp,encoding 192 amino acids.The protein molecular formula encoded by the gene was C₉₆₄H₁₅₂₅N₂₇₅O₂₉₁S₈,the theoretical isoelectric point of the protein was 8.91,the instability coefficient and the total average hydrophilicity were 84.14 and -0.822,respectively,which belonged to the unstable hydrophilic protein.There were 29 potential phosphorylation sites,including 14 serines,11 threonines and 4 tyrosines.There was no signal peptide and transmembrane helix structure.Subcellular localization showed that it mainly existed in the nucleus and belonged to non-secretory protein.The high-level structure of the protein was mainly composed of 39.06% α-helix and 56.25% random coil structure.Homology analysis showed that the genetic relationship between yak and cattle was the closest,which was in line with the actual situation,indicating that the Akirin1 gene was relatively conservative in the evolution process.Real-time Fluorescence Quantitative results showed that Akirin1 gene was expressed in heart,liver,spleen,lung,kidney,muscle and adipose tissue of adult Gannan yak,and the relative expression in kidney tissue was the highest,which was significantly higher than that in other tissues,and the expression in muscle and liver tissue was relatively low.This experiment provides a theoretical basis for further study on the functional characteristics of yak Akirin1 gene.

Key words: Yak, Akirin1 gene, Cloning, Bioinformatic analysis, Tissue expression nalysis