与人体肥胖相关的神经肽Y受体Y1基因的克隆及序列分析

Cloning and sequence analysis of the human obesity - related neu- ropeptide Y （NPY） receptor Y1 gene

目的克隆与肥胖相关的人神经肽Y受体Y1（NPY1R）基因，并鉴定该克隆基因序列的正确性。方法从人的脂肪组织中提取总RNA并进行反转录，以此为模板用PCR扩增NPY1R基因的eDNA，然后与pET28a＋载体重组，筛选阳性克隆和DNA序列分析鉴定，测序后与GeneBank中NPY1R基因进行同源性比较和序列分析。结果PCR扩增出一个1100bp左右的DNA片段，与载体重组后DNA序列分析鉴定，DNA序列分析显示克隆的DNA片段是人NPY1R基因，且所克隆的基因共编码384个氨基酸，分子量为44KD，与GeneBank中NPY1R基因序列同源性达100％。结论克隆的人NPY1R基因与GeneBank中NPY1R序列完全一致，为进一步应用分子生物学技术深入研究NPY1R与人体肥胖发生、发展及转化等相关作用机制及应用该基因进行其基因蛋白表达奠定了基础。

Objective To clone the human neuropeptide Y receptor Y1 （ NPY1 R）, which is one of the human obesity - related genes, and identify the cloned gene sequence. Methods The eDNA of NPY1R gene was amplified from RNA of the human fat by reverse transcription polymerase chain reaction （ RT - PCR） . The PCR products were recombined with pET28a ＋ vector, the positive clones were selected and DNA sequence analysis was conducted for the identification, followed by sequence analysis and homology comparison with that reported in Genebank after sequencing. Results A DNA fragment of about 1100bp was obtained by RT - PCR, the eDNA se- quence of the recombinant plasmid of pET28a ＋ with the fragment was analyzed, and the result showed that the cloned DNA fragment was just human NPY1R eDNA. The cloned gene encoded 384 amino acids with a molecular weight of 44KD. The nucleotide sequence homology of the cloned NPY1R was 100% in comparison with that reported in Genebank. Conclusions The sequence of NPY1R eDNA cloned successfully is the same as that of NPY1R in Genebank, which lays the foundation for further studies of gene expression of NPY1R and the interrelationship between NPY1R and genesis, development and transformation of obesity by means of molecular biology.