摘要
目的克隆大鼠肝再生增强因子 (ALR)编码序列 ,并在原核细胞表达。方法按文献报道大鼠肝再生增强因子核苷酸序列合成引物 ,从2周龄大鼠肝组织提取RNA ,利用RT_PCR扩增大鼠肝再生增强因子基因编码区 ;将PCR扩增片断亚克隆到pBV221质粒 ,构建原核表达重组载体 ,导入到大肠杆菌后热诱导表达目的蛋白。结果从大鼠肝脏的RNA中扩增到长约400bp的ALRcDNA编码区基因 ,序列分析表明与文献报道一致 ;重组克隆经热诱导表达出分子量约15kD大小的蛋白质 ,与预期值一致。结论从2周龄大鼠肝组织中成功克隆肝再生增强因子基因 。
Objective To clone the gene of rat augmenter of liver regeneration (ALR), and express the protein encoded by ALR in the prokaryotic cells. Methods The coding region of ALR gene was amplified by a coupled one_step reverse transcription PCR (RT_PCR). The PCR fragment of ALR gene was subcloned to pBV22l plasmid, and constructed a prokaryotic expression vector, and then the recombinant plasmid was heat_induced to express protein after transducted to E.coli. Results ALR cDNA of 400bp was amplified successfully, which was consistent with the gene reported in literature after nucleotid sequence was detected. The specific expression band was observed in SDS_PAGE analysis, and its moleculor weight was about 15 kD which was in accordance with the expecting value. Conclusions The augementer of liver regeneration gene was cloned from the liver tissue of 2_week_old SD rat and the protein encoded by the gene was expressed in the prokaryotic cells.
