人RANTES基因克隆与测序

Cloning and Sequencing of Human RANTES Gene

目的 获得人趋化因子RANTES基因,用于抗艾滋病转基因中药的培育。方法 RT-PCR扩增、目的基因片段与T-载体连接和有用基因的序列分析。结果 通过淋巴细胞激活、RNA分离和RT-PCR扩增,已获得预期的276bp片段,并经PCR和酶切反应进行了鉴定。初步测序结果表明,克隆基因片段包含了编码RANTES基因信号肽与成熟蛋白的完整序列以及起始和终止密码,但编码信号肽的-10位密码子(GTT)与已发表的序列(GCT)不同。这一差异是来自基因本身的突变抑或为实验误差尚有待进一步查明。结论 克隆了人RANTES基因并进行了测序。

Objective To obtain human chemokine RANTES gene for the cultivation of a transgenic herbal medicine a-gainst AIDS. Methods RT- PCR amplification, ligation of the target fragment with a T- vector and sequence analysis of the interested gene were employed. Results An expected 276bp fragment was amplified by RT-PCR from RNA of activated lymphocyte, and identified by both PCR and restriction enzyme digestion. The preliminary sequence data shown that the cloned fragment comprised the complete sequence encoding its signal peptide and mature protein of RANTES, as well as the start and end codons. The-10 codon (GTT) for signal peptide, however, was different from the published sequence (GCT). Whether this variation is derived from mutation of the gene itself or due to an experimental error needs to be further elucidated. Conclusion The RANTES gene of human being has been cloned and sequenced in China.