鸡Smad4基因克隆、生物信息学及组织表达分析

试验旨在克隆鸡Smad4基因,并对其进行生物信息学和组织表达分析。以苏禽3号优质黄羽肉鸡为试验对象,使用RACE技术扩增并克隆了鸡Smad4基因全长序列,对其进行生物信息学分析,构建系统进化树,并利用实时荧光定量PCR检测了Smad4基因在鸡各组织中的表达情况。结果显示,鸡Smad4基因全长序列包含17 bp的5'UTR、1 842 bp的开放阅读框和466 bp的3'UTR,有11个外显子和10个内含子,mRNA序列全长2 325 bp,可编码613个氨基酸。系统进化树显示,鸡Smad4与其他鸟类聚为一类。生物信息学分析显示,Smad4蛋白分子质量为65.43 ku,理论等电点为10.04,为亲水蛋白;含有2个保守结构域：MH1和MH2;二级结构由α-螺旋（18.11%）、延伸链（17.29%）和无规则卷曲（64.60%）组成。组织表达分析表明,Smad4基因在鸡的心脏、肝脏、空肠、卵巢中均有较高表达,而在胸肌中不表达。本试验成功获得了鸡Smad4基因全长序列,并初步研究了其组织表达规律,为进一步研究Smad4基因在鸡繁殖活动中的分子机制提供了理论依据。

Cloning,Bioinformatics and Tissue Expression Analysis of Smad4 Gene in Chicken (Gallus gallus)

The purpose of this study was to clone Smad4 gene and analyze its bioinformatics and tissue expression in chicken. Suqin No. 3 chicken was selected as experimental subjects, the full-length sequence of Smad4 gene in chicken was amplified by Race technology and cloned, the bioinformatics analysis of Smad4 gene was carried out, and the phylogenetic tree was constructed. The expression of Smad4 gene in chicken tissues was detected by Real-time quantitative PCR. The results showed that a full-length of Smad4 gene in chicken was obtained, which contained 17 bp 5'UTR, 1 842 bp open reading frame and 466 bp 3'UTR. There were 11 exons and 10 introns, the length of mRNA was 2 325 bp, which encoded 613 amino acids. The phylogenetic tree showed that Smad4 gene in chicken was clustered with other birds. Bioinformatics analysis showed that the molecular mass of Smad4 protein was 65.43 ku, the theoretical isoelectric point was 10.04, it was a hydrophilic protein. Smad4 protein contained two conserved domains MH1 and MH2. The secondary structure was consist of alpha helix (18.11%), extended chain (17.29%) and random coil (64.60%). Tissue expression analysis showed that Smad4 gene was highly expressed in heart, liver, jejunum and ovary, but there was no expression in breast muscle. In this study, the full-length sequence of Smad4 gene in chicken was successfully obtained, and its tissue expression pattern was preliminarily studied, which provided a theoretical basis for further study on the molecular mechanism of Smad4 gene in reproductive activity of chicken.