人乳铁蛋白基因克隆及表达载体构建

Cloning of human lactoferritin gene and construction of expression vector

通过 PCR法扩增出 2 366bp的人乳铁蛋白 c DNA、80 0 bp的山羊β -乳球蛋白基因 5′端调控序列 ,经 PCR反应连接后 ,克隆到表达载体 p LNCX中 .测序结果表明 :与 Gene Bank中登录的序列相比 ,所获人乳铁蛋白 c DNA序列的同源性为 99.74 % ,山羊β -乳球蛋白基因 5′端调控序列的同源性为 99.75% .酶切结果证明得到了正确的重组载体 ,可用于乳腺生物反应器的研究 .利用 PCR反应直接克隆目的片段 ,并通过同源序列进行 PCR连接 ,极大提高了克隆的速度和准确性 ,为基因克隆及其表达研究带来了方便 .

The 2 366 bp cDNA of human lactoferritin and 800 bp goat β lactoglobulin 5′ flanking regulatory sequence are obtained by PCR method. These two sequences were connected by PCR and then were cloned to expression vector pLNCX. The homology of human lactoferritin cDNA sequence and goat β lactoglobulin are 99.74% and 99.75% respectively comparing with those of Gene Bank. The results of enzyme digest indicate that the correctly reconstructive vector is obtained. The authors directly cloned the target fragments and connected them by PCR, therefore improved the velocity and the accuracy of cloning. This method is convenient to study gene expression.