人睫状神经营养因子基因的克隆及序列分析

Clone and Sequence Analysis of Ciliary Neurotrophic Factor Gene

目的:构建1例2型糖尿病足坏疽截肢患者睫状神经营养因子(hCNTF)cDNA的克隆,为进一步研究睫状神经营养因子与2型糖尿病及其并发症发病的关系奠定基础。方法:从糖尿病足坏疽截肢患者坐骨神经组织中提取总RNA,经逆转录-聚合酶链式反应(RT-PCR)扩增hCNTFcDNA完全编码区,扩增产物经纯化后克隆至pMD18-T载体,并进行测序及序列分析。结果:成功构建此患者CNTFcDNA的克隆,重组质粒(pMD18-T/CNTF)经测序与Genebank检索的hCNTFcDNA突变序列(AccessionNM_S72921)100%符合。结论:构建的突变CNTFcDNA克隆为进一步研究CNTF与2型糖尿病及其并发症在发病上的关系提供线索。

Objective： To set out the foundation for further research of the pathogenic relationship between ciliary neurotrophic factor （CNTF） and type 2 diabetes （T2DM） by cloning the cDNA of human CNTF from a diabetic foot wound. Methods： To extract total RNA of human sciatic nerve tissue from a diabetic patients with foot wound and amplify the cDNA of human CNTF by RT-PCR. The amplified product was purified and cloned into plasmid pMD-18T. The cloned nucleotide was sequenced and analyzed. Results： The cloning of cDNA of human CNTF was successful. The sequence shared 100% homology with that of Genebank （Accession NM_S72921）. Conclusion： The constructed clone of mutated cDNA of human CNTF showed that there would be a possible relationship between CNTF and diabetic complications.