转生长激素基因中华倒刺鲃的构建及其检测

The Construction and Detecting of Transgenic Spinibarbus sinensis With Growth Hormone Gene

用从斑点叉尾鮰(Ictalurus punctatus)cDNA文库克隆到的生长激素基因,与鲤β肌蛋白启动子、大麻哈鱼生长激素基因的poly(A)终止信号序列等调控元件共同构建了表达重组质粒pFV2 cfGH。提取重组质粒,用精子载体法导入中华倒刺鲃(Spinibarbus sinensis)的受精卵中,经孵化培养成转基因成鱼。抽提423尾转基因实验鱼血总DNA,经PCR检测表明:斑点叉尾鮰生长激素基因已导入中华倒刺鲃基因组中,导入率为17.26%。同时通过RT-PCR技术检测阳性中华倒刺鲃中生长激素基因的表达情况。

The expression recombinant plasmid pFV2cfGH was constructed from growth hormone gene which was cloned from the channel catfish cDNA library, the β - action promoter of carp and the polyadenylation signal from the salmon growth hormone gene, and other controlling elements. The recombinant plasmid pFV2ctGH was transmitted into the fertilized eggs of the Spinibarbus sinensis through Sperm - mediated gene transfer, and was hatched into transgenic fish. Total DNA of 423 transgenic Spinibarbus sinensis was abstracted, and growth hormone gene was detected by PCR. The results showed that the growth hormone gene of the channelfish had been transmitted into the genome of the Spinibarbus sinensis and the integration rate is 17.26%. At the same time, the expression of growth hormone gene in positive Spinibarbus sinensis was detected by RT - RCR.