人骨形态发生蛋白4成熟肽cDNA的克隆及测序

背景：研究表明骨形态发生蛋白4的骨诱导能力最强,但在组织中含量甚微。目的：克隆人骨形态发生蛋白4成熟肽基因。方法：从人胎盘组织中提取信使核糖核酸,根据基因库人骨形态发生蛋白4基因序列合成两条引物,利用一步法反转录聚合酶链式反应技术扩增出人骨形态发生蛋白4成熟肽的基因序列,将聚合酶链反应扩增产物基因片段连接克隆载体质粒中转化大肠杆菌DH5α进行克隆筛选鉴定及测序。结果与结论：琼脂糖凝胶电泳检测反转录聚合酶链式反应产物及阳性克隆质粒经双酶切产物显示一长约370bp的条带,脱氧核糖核酸测序结果与基因库中的序列相符。结果证实,利用反转录聚合酶链式反应技术可成功的从人胎盘组织中克隆出人骨形态发生蛋白4成熟肽基因。

Cloning and sequencing of mature peptiede of human bone morphogenetic protein 4 cDNA

BACKGROUND：Studies have shown that bone morphogenetic protein 4（BMP-4） has strongest osteoinductive capacity,but its content in tissue is low.OBJECTIVE：To clone human BMP-4（hBMP-4） mature peptide gene.METHODS：According to the hBMP-4 sequence reported in Genebank,two primers were designed.The gene sequence of hBMP-4 mature peptide was gained by one step RT-PCR from the mRNA,which was extracted from human placenta.Then it was cloned into the vector of PGEM-T and the resultant plasmid was transformed into DH5α.The sequence of the plasmid BMP-4 was detected by DNA sequence machine.RESULTS AND CONCLUSION：DNA agarose electrophoresis showed that the product of RT-PCR was about 370 bp and it was consistent with the reported hBMP-4 sequence in Genebank.These findings suggest that the mature peptide of hBMP-4 gene can be successfully cloned from human placenta by RT-PCR.