藏羊GM-CSF基因的RT-PCR扩增、序列分析及蛋白结构预测  

RT-PCR Amplification and Sequence Analysis of GM-CSF Gene of Tibetan Sheep and Protein Structure Prediction

作　　者：张晓芬[1] 冶贵生[1] 马玉花[1] 贺晓龙[1] 康明[1] 韩志辉[1] 贾跃宁 张洪波[1] 

机构地区：[1]青海大学农牧学院,省部共建三江源生态与高原农牧业国家重点实验室,西宁810016

出　　处：《基因组学与应用生物学》2016年第10期2662-2667,共6页Genomics and Applied Biology

基　　金：青海省科技厅项目(2016-ZJ-738);青海大学“123高层次人才培养工程”项目共同资助

摘　　要：为了研究藏羊GM-CSF基因及其编码蛋白的功能,提取藏羊脾脏总RNA,应用RT-PCR技术对藏羊GM-CSF基因进行扩增及测序,并利用DNA Star软件进行序列分析及编码蛋白结构预测。测试表明,藏羊GM-CSF基因长度为381 bp,编码127个氨基酸;藏羊GM-CSF基因与参考绵羊、藏羚羊、山羊和水牛的GM-CSF基因的核苷酸序列同源性依次为100%、99.2%、98.7%和92.1%,氨基酸序列同源性依次为100%、97.6%、96.9%和81.0%。蛋白结构预测表明GM-CSF蛋白具有1个N-糖基化位点、1个N-豆蔻酰化位点、1个粒细胞-巨噬细胞集落刺激因子信号、2个蛋白激酶C磷酸化位点、3个酪蛋白激酶Ⅱ磷酸化位点;综合二、三级结构预测得出,该蛋白主要由α螺旋组成,其次是β转角和无规则卷曲,而β折叠相对较少。研究成果可为青海藏羊GM-CSF基因抗病功能的研究提供参考。In order to study the function of GM-CSF gene and its encoding protein of Tibetan sheep, the total RNA was firstly extracted from spleen of Tibetan sheep, then the GM-CSF gene of Tibetan sheep was amplified and sequenced by RT-PCR. Finally, the gene sequence and protein structure of GM-CSF were analyzed and predicted by DNA Star software. The results showed that the length of GM-CSF gene was 381 bp, and it encoded 127 amino acids. Compared with GM-CSF gene of ovis aries, pantholops hodgsonii, capra hircus and bubalus bubalis, the nucleotide sequence homology was 100%, 99.2%, 98.7% and 92.1% respectively, and the amino acid sequence homology was 100%, 97.6%, 96.9% and 81.0% respectively. The results of protein prediction showed that the GM-CSF protein had one N-glycosylation site, one N-myristoylation site, one granulocyte-macrophage colony-sti mulating factor signature, two protein kinase C phosphorylation sites and three casein kinase Ⅱ phosphorylation sites, besides, the results of protein secondary and tertiary structure prediction showed that the GM-CSF protein consisted mainly of β-helix, followed by β-turn and random coil, and B-pleated sheet was relatively less. These results can provide references for studying the resistant function of GM-CSF gene of Qinghai Tibetan sheep.

关 键 词：藏羊 GM-CSF基因 RT-PCR 序列分析 蛋白结构预测 

分 类 号：S826[农业科学—畜牧学]