西藏小型猪IGF-1基因的克隆测序分析

Analysis of the cloned IGF-1 gene cDNA sequence of Tibet minpig

目的克隆西藏小型猪的肝脏组织中的IGF-1基因的c DNA序列,并与Pubmed中查询到的猪c DNA进行比对分析。方法提取了西藏小型猪肝脏组织的总RNA,应用RT-PCR技术扩增了IGF-1基因的c DNA序列,将扩增出的片段克隆到p MD18-T载体上,构建重组质粒p MD18-T-IGF-1,进行测序分析。结果克隆出西藏小型猪肝脏组织中的IGF-1的c DNA序列,获得了大小为567 bp长的片段,编码了186个氨基酸,与Pubmed中查询到的猪(NM_214256.1)的IGF-1基因高度同源,比对序列发现,在440、455 bp处发生了G→A、C→T的突变,该位点的突变引起相应编码氨基酸的变化,分别由组氨酸变成了精氨酸、亮氨酸转变成了丝氨酸。结论为西藏小型猪的生长发育机制研究提供了分子学依据。两个位点的突变引起的氨基酸的改变是否是导致西藏小型猪矮小的原因,需要进一步论证。

Objective To compare and analyze the sequence between Tibet minipigs and Pubmed pig （NM_ 214256. 1 ） IGF-1 eDNA by cloning and sequencing the IGF-1 gene eDNA of the liver tissue of Tibet minipigs. Methods Total RNA was extracted from Tibet minipig liver and amplified the sequence of IGF-1 gene by RT-PCR. The amplified fragment was cloned into pMD18-T vector, and the recombinant plasmid pMD18-T-IGF-1 was constructed and sequenced. Results Tibet minipig IGF-1 gene fragment containing the coding region of 567 bp. The coding region encoded 186 amino acids, and 99% were homologous compared with that of the pig （ NM_214256. 1 ）. Compared with pig （ NM_214256. 1 ） , we found that the mutation in the 440 bp, 455 pb, occurred at G→A, C→T mutation, and the corresponding amino acids were changed, histidine→arginine and leueine→serine, respectively. Conclusions The results of this study provide a molecular basis for breeding research. Whether the mutation at the two sites changes the protein function and body size or not need to be further studied.