荷包猪DR基因座α链和β链基因的扩增与序列分析
发布时间：2018-09-12 05:48
【摘要】：荷包猪属于我国地方优质品种猪,是经过原始进化而成的物种,具有重要的科学研究价值。猪的DR基因座属于猪主要组织相容性复合体(maj or histocompatibility complex,MHC),亦即猪白细胞抗原(swine leukocyte antigen,SLA)II类分子重要功能基因,包括α链(DRA)和p链(DRB),是物种分子进化的标记。为研究荷包猪SLA-DR基因,以荷包猪脾脏组织为材料,采用RT-PCR技术,分别以引物DRAF1/DRAR1扩增sLA-DRA、以引物DRBF1/DRBR1扩增sLA-DRB。扩增产物分别与pMD18-T载体连接,并转化宿主菌JM109。EcoR I和Hind Ⅲ双酶切筛选阳性克隆。最后利用生物软件DNAMAN和Mega 5等进行比对、分析及构建分子进化树。结果显示,成功克隆3个荷包猪SLA.DRA.HB01-03和3个SLA.DRB.HB01-03等位基因。序列测定结果显示,α链3个等位基因SLA.DRA.HB0l-03长度为779 bp,编码252个氨基酸,分别获得序列号LC002668.LC002669和LC002670；p链3个等位基因SLA-DRB-HB01-03为836bp,编码266个氨基酸,分别获得序列号LC20542、LC20543和LC20544。氨基酸序列对比分析发现,α链等位基因SLA-DRA.HB01.03与其他SLA-DRA等位基因相比较,变异位点较少。对于p链基因SLA-DRA-HB01-03.与其他SLA-DRB等位基因相比较变异位点较多。一致性分析显示,SLA-DRA-HB01-03与其他SLA-DRA等位基因一致性为98.7%-99.6%,而SLA-DRB-HB01-03与其他SLA-DRB等位基因一致性为90.3%-99.8%。分子进化树分析结果显示,SLA-DRA-HB01-03自成一体系,但与其他等位基因的距离均较近；而荷包猪SLA-DRB-HB01-03进化树也自成一系,与其他等位基因相比较进化距离较远,进化更加原始。本研究成功克隆了荷包猪DR基因座α链和p链,证明荷包猪DR基因座α链基因比较保守,而β链基因多态性较丰富,进化上更具有特征性。本研究为今后进一步研究荷包猪SLA-DR基因座α链和β链基因结构和功能奠定基础。
[Abstract]:The purse pig belongs to the local high quality pig of our country. It is a species of primitive evolution and has important scientific research value. Porcine DR loci belong to the porcine major histocompatibility complex (maj or histocompatibility complex,MHC), that is, porcine leukocyte antigen (swine leukocyte antigen,SLA) II class important functional genes, including 伪 -chain (DRA) and p-chain (DRB), is the molecular evolution of species markers. In order to study the SLA-DR gene of the purse pig, the spleen tissue of the purse pig was used as the material, the sLA-DRA, was amplified by the primers DRAF1/DRAR1 and the sLA-DRB. was amplified by the primer DRBF1/DRBR1 using RT-PCR technique. The amplified products were ligated with pMD18-T vector respectively, and the positive clones were screened by double digestion of JM109.EcoR I and Hind 鈪，
本文编号：2238088
[Abstract]:The purse pig belongs to the local high quality pig of our country. It is a species of primitive evolution and has important scientific research value. Porcine DR loci belong to the porcine major histocompatibility complex (maj or histocompatibility complex,MHC), that is, porcine leukocyte antigen (swine leukocyte antigen,SLA) II class important functional genes, including 伪 -chain (DRA) and p-chain (DRB), is the molecular evolution of species markers. In order to study the SLA-DR gene of the purse pig, the spleen tissue of the purse pig was used as the material, the sLA-DRA, was amplified by the primers DRAF1/DRAR1 and the sLA-DRB. was amplified by the primer DRBF1/DRBR1 using RT-PCR technique. The amplified products were ligated with pMD18-T vector respectively, and the positive clones were screened by double digestion of JM109.EcoR I and Hind 鈪，
本文编号：2238088
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