人粒细胞集落刺激因子cDNA的克隆、序列鉴定及初步表达

Molecular Cloning and Sequencing of the cDNA for Human G-CSF and its Preliminary Expression

利用多聚酶链反应技术,从人膀胱癌细胞系5637细胞中快速扩增并克隆了人粒细胞集落刺激因子cDNA,序列分析证明,该cDNA包含人粒细胞集落刺激因子的全部编码基因,全长612bp,编码30个氨基酸的信号肽和174个氨基酸的成熟蛋白。其中第43位codon出现一个碱基的突变(CAC→TAC)导至第43位氨基酸的改变(组氨酸→酪氨酸)。经逆转录病毒导入SP2/0细胞并初步表达。结果表明:该基因产物具有G-CSF活性。

The cDNA for human G-CSF was cloned from total RNA of human bladder carcinoma 5637 cells following PCR amplification. Sequencing data proved that the cloned cDNA contained the intact coding region of human G-CSF gene which was 612 bp in length, encoding 30 amino acids of signal peptide and 174 amino acids of the mature protein. One base mutation was found in the 43rd codon of the cDNA (CAC→TAC) resulting in the replacement of histidine by tyrosine. The gene products proved having G-CSF aetivity after its preliminary expression in SP2/0 cells following retroviral transfer.