%0 Journal Article
%T 犬前松弛素原基因的克隆与生物信息学分析
%A 任志华
%A 夏娟
%A 刘欢欢
%A 余树民
%A 沈留红
%A 曹随忠
%A 左之才
%A 邓俊良
%A 马晓平
%J 南京农业大学学报
%D 2016
%R 10.7685/jnau.201509004
%X [目的]本试验通过对犬前松弛素原基因的分析推导获得犬松弛素基因序列，为犬松弛素多克隆抗体的制备及临床应用提供理论基础。[方法]以北京犬和吉娃娃新生胎盘、贵宾犬卵巢为材料，分别提取总RNA，经RT-PCR扩增，获得犬前松弛素原基因，应用生物信息学方法初步分析犬前松弛素原基因的结构及功能。[结果]完整克隆了从所获样品中扩增到的预期大小目的片段，包含一个534 bp的开放阅读框，编码177个氨基酸。扩增片段序列分析结果显示，与NCBI核酸数据库检索的犬前松弛素原的cDNA序列同源性为99.6%，均有3个核苷酸位点发生改变，从而引起相应氨基酸改变，但并未引起蛋白结构的变化。进化树分析结果显示，所获3条目的片段同为一支，与猫前松弛素原基因的进化距离最近。生物信息分析显示，3条序列等电点、带电荷数、最高（最低）氨基酸、消光系数、半衰期、不稳定系数均一致，相对分子质量、不稳定系数、脂肪系数、总平均疏水指数有细微差异；跨膜区、信号肽、糖基化位点分析结果一致；二级结构组成一致，包含α-螺旋、延伸链、无规则卷曲，比例存在细微差异。[结论]成功克隆了犬前松弛素原基因，可以生物信息学分析为基础进行犬松弛素多克隆抗体的制备。</br>[Objectives] The aim of our study was to know the gene sequence of relaxin gene by cloning and bioinformatics analysis of the preprorelaxin gene of canine. [Methods] This study was based on the Pekingese and the newborn Chihuahua’s placenta and the Poodle ovary,total RNA was extracted and amplified by RT-PCR,and then respectively,the preprorelaxin gene was obtained,and preliminary analysis of the structure and function of the preprorelaxin gene of the dog was conducted by using bioinformatics method. [Results] Results showed that the experiment completely cloned the samples obtained from the expansion to the expected purpose fragment size,containing an open reading frame of 534 bp,encoding 177 amino acids. Sequence analysis of the amplified fragments showed that the percentage of the homology of the cDNA sequence of preprorelaxin with NCBI nucleic acid database was 99.6%,having three nucleotide site changes,so as to cause the corresponding amino acid changes,but did not cause changes in the structure of the protein. Phylogenetic tree analysis results showed that,the three target fragments were the same as one,and the evolutionary distance of the cat preprorelaxin gene was nearest. Bioinformatics analysis showed that,the 3 sequences were the same in the electric point,the charge number,the highest minimum amino acid,the extinction coefficient,the half-life and the instability coefficient. The molecular weight,the coefficient of instability,the fat coefficient,and the total average hydrophobicity index had slight differences. The crossing membrane region,signal peptide and glycosylation site analysis results were consistent. And the secondary structure was the same including a-helix,an extended chain,a non regular,and having a slight difference. [Conclusions] We successfully cloned the preprorelaxation gene of canine. And the polyclonal antibody against canine can be prepared depending on the bioinformatics analysis
%K 犬
%K 前松弛素原基因
%K 松弛素
%K 生物信息学分析&lt
%K /br&gt
%K canine
%K preprorelaxin gene
%K clone
%K bioinformatics analysis
%U http://nauxb.njau.edu.cn/oa/darticle.aspx?type=view&id=201606021