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目的:研究湖南大围子猪SLA-DR基因特性,评价该猪种在异种器官移植中是否具有应用前景。方法:应用RT-PCR扩增大围子猪SLA-DRA和SLA-DRB基因,插入pUCM-T载体,双向测序,用NCBI中的BLAST和ExPASY软件进行生物信息学分析。结果:大围子猪SLA-DRA和SLA-DRB基因扩增片段大小分别为1177bp和909bp,均包含完整的开放阅读框,分别编码252和266个氨基酸残基。生物信息学分析结果表明,大围子猪SLA-DRA和SLA-DRB与人类相应的DRA、DRB相比,氨基酸同源性分别为82%和73%。SLADRα链与人CD4分子结合部位介于第124至136位氨基酸,大围子猪SLA-DRA在该结合区域与人类相应的DRA存在两个氨基酸差异,即:第127位(lle→Val),第136位(Ser→Thr);SLADRβ链与人CD4分子结合部位位于第134至148位氨基酸,大围子猪SLA-DRB在该结合区域与人类相应的DRB相比氨基酸序列完全相同。与国际GenBank已登录的许多猪种相比,SLA-DRA基因同源性高达100%,而SLA-DRB基因在各猪种之间具有很高的多态性。结论:克隆湖南大围子猪SLA-DRA和SLA-DRB基因,该基因与人类相应的HLA-DRA和HLA-DRB基因高度同源,提示该猪种有望作为异种移植的候选供体。
OBJECTIVE: To study the SLA-DR gene characteristics in Daweizi pigs in Hunan province, and to evaluate whether this pig breed has potential application in xenotransplantation. Methods: The gene of SLA-DRA and SLA-DRB of Daweizi pig was amplified by RT-PCR and inserted into pUCM-T vector. The two vectors were sequenced and the bioinformatics analysis was carried out by using BLAST and ExPASY software in NCBI. Results: The size of amplified fragments of SLA-DRA and SLA-DRB in Daweizi pigs were 1177bp and 909bp, respectively, and contained complete open reading frames (ORFs) encoding 252 and 266 amino acid residues respectively. The results of bioinformatics analysis showed that the amino acid homologies of SLA-DRA and SLA-DRB in Daweizi pigs were 82% and 73% compared with those of human corresponding DRA and DRB, respectively. SLADRα chain binds to human CD4 molecule between amino acids 124 to 136. There is two amino acid differences in SLA-DRA between human and overexpressing DAA in the binding region, namely, the 127th (lle → Val), the 136 (Ser → Thr); the SLADRβ chain binds to amino acids 134 to 148 of the human CD4 molecule, and the megalobas SLA-DRB has identical amino acid sequences in this binding region to the human corresponding DRB. The SLA-DRA gene homology is as high as 100% compared with many pigs registered with GenBank in the world, while the SLA-DRB gene has a high polymorphism among the pig breeds. CONCLUSION: The SLA-DRA and SLA-DRB genes were cloned from Hunan Daweizi pigs. This gene is highly homologous to the corresponding human HLA-DRA and HLA-DRB genes, suggesting that this pig breed is expected to be a candidate donor for xenotransplantation.