[目的]克隆广西德保猪抵抗素(Resistin,RENT)基因并进行系统生物信息学分析,阐明德保猪脂肪细胞分化与脂肪生成的分子机制.[方法]根据NCBI已公布的猪RENT基因序列(NM 213783.2)设计特异性引物,以从德保猪血液中提取的总RNA为模板,PCR扩增RENT基因序列,T-A克隆至pEASY-T1载体后进行测序分析,并将所得测序结果用BLAST程序、MEGA 5.0软件、EXPASY服务器、SMART程序、SignIP程序、Softberry服务器和DNASTAR软件进行生物信息学分析.[结果]扩增获得468 bp的RENT基因序列片段,包含全长的德保猪RENT基因编码区和部分非编码区序列.多重序列比较分析结果显示,德保猪RENT基因编码区核酸序列与猪、牛、山羊、绵羊、人、兔、猕猴和豚鼠的同源性分别为100.0%、85.5%、85.5%、84.5%、81.2%、80.3%、80.0%和77.3%.德保猪RENT蛋白结构预测结果显示,其理论分子质量11.69 ku,等电点7.82,为弱碱性蛋白;N-末端存在信号肽,定位于胞质外,仅具有1个低复杂度结构域,包含1个α-螺旋、8个β-折叠、7个T-转角及两个无规则卷曲.[结论]RENT基因在德保猪中呈高度保守特征,是其脂肪细胞分化中的重要调控基因.
[Objective] Cloning and bioinformatics analysis of Resistin gene in Debao pig were conducted to explore molecular mechanism of adipocyte differentiation and lipogenesis of Debao pig.[Method]With the released pig RENT gene sequence(NM_213783.2) as a template,a pair of primers was designed.The RENT gene was amplified by RTPCR from isolated total RNA of Debao pig blood.The RENT gene was inserted into pEASY-T1 vector by T-A clone method,and was sequenced.And its bioinformatics analysis was performed using BLAST program,MEGA5.0 software,EXPASY server,SMART program,SignIP program,Softberry server and DNAStar program.[Result]A 468bp RENT gene fragment of Debao pig was cloned and sequenced.The sequence multi-aligned results showed that Debao pig RENT gene shared 100.0%,85.5%,85.5%,84.5%,81.2%,80.3%,80.0% and 77.3% homology with that of Sus scrofa,Bos taurus,Capra hircus,Ovis aries,Homo sapiens,Oryctolagus cuniculus,Macaca mulatta and Cavia porcellus,respectively.The further protein structure bioinformatics analysis suggested that it was a weakly basic protein with the weight of 11.69 ku and isoelectric point of 7.82.It had an N-terminal signal peptide,localized in the extranuclear with a low complexity domain containing 1α-helices,8 β-folds,7 T-corners and 2 random coils.[Conclusion]Resistin gene fragment in Debao pig is highly conserved in multiple species,and it is of great importance in controlling adipose cell differentiation of Debao pig.