摘要
根据已发表的IL-18蛋白cDNA序列保守区设计一对特异性引物,应用RT-PCR技术从ConA活化的牛外周血单核细胞中扩增到编码牛IL-18蛋白基因,其大小为582 bp。将该基因克隆到pMD18-T载体中,经序列测定表明该基因与gengbank上发表的牛白细胞介素-18基因序列同源性为98%。将该基因从重组pMD-gIL-18质粒中亚克隆到表达载体pET-32 a(+)质粒中,构建原核表达重组质粒,经酶切和PCR鉴定后表明构建的重组质粒为阳性。
According to the encoding sequence of the IL-18 protein and the multiple cloning sites characteristic of the prokaryotic expression vector pET-32 a （＋） in Genebank, appropriate primers specific to IL-18 gene were designed. The target DNA fragments were obtained by RT-PCR amplification and then were cloned into the pMD18-T vector. After identification by restriction enzyme digestion and sequencing analysis, the specific DNA fragments were cloned into the prokaryotic expression vector pET-32 a （＋）. The positive recombinant plasmids were named pET-32a （＋）-IL-18 after identification by restriction enzyme digestion and PCR amplification. Results showed that recombinant expression plasmids pET-32a（＋）-IL-18 were successfully constructed.
出处
《黑龙江八一农垦大学学报》
2009年第6期51-53,共3页
journal of heilongjiang bayi agricultural university
关键词
白细胞介素
原核表达
载体构建
IL-18
prokaryotic expression
vector construction
