人骨形态发生蛋白-4成熟肽cDNA的克隆

Cloning and sequence of mature peptide of human bone morphognetic protein-4 cDNA

目的 :克隆人骨形态发生蛋白 4完整成熟肽的基因。方法 :根据Genebank人骨形态发生蛋白 4的基因序列化学合成两条引物 ,从人胎盘组织中提取得到总RNA ,用反转录聚合酶链式反应 (RT PCR )技术扩增出长 0 .3 4kb编码人骨形态发生蛋白 4(BMP 4)成熟肽的基因序列。将所得的基因片段插入克隆载体PBluescriptⅡSK质粒中并转化大肠杆菌TG 1,提取重组质粒 ,酶切鉴定并测序。结果 :RT PCR产物琼脂糖凝胶电泳显示一长约 3 70bp的条带 ,阳性克隆质粒经双酶切可切出约 3 5 0bp的片段 ,全自动DNA测序结果表明和Genebank中的序列完全相符。结论 :通过反转录聚合酶链式反应可从人胎盘组织中成功克隆出人BMP 4成熟肽基因。

Objective:To clone the cDNA of mature peptide of human bone morphogenetic protein 4(hBMP 4).Methods: Two primers were designed according to the hBMP 4 sequence reported in Genebank.The mature peptide of hBMP 4 cDNA was gained by RT PCR from the mRNA,which was extracted from human placenta.Then it was cloned into the vector of Pbluescript Ⅱ SK plasmid and the sequence of the plasmid Pbluescript Ⅱ SK BMP 4 was detected by DNA sequence machine.Results: DNA agarose gel electrophoresis showed that the product of RT PCR was about 370bp,it was corresponed with the result of two enzyme digestion of the recombinanted plasmid.The result of sequence assay was in agreement with the reported hBMP 4 sequence in Genebank.Conclusion:The mature peptide of hBMP 4 cDNA was cloned successfully and the sequence was testified correctly.