摘要
利用高保真PCR法,分别扩增了牛乳腺β-酪蛋白基因的1.8和1.1 kb的5′和3′调控序列,将其分别克隆入TA载体。经PCR验证后测序,用NCBI Blast软件分析表明其克隆片断与奶牛β-酪蛋白基因相应区域同源性分别为97.0%和99.0%,表明成功克隆了酪蛋白基因5′和3′的调控区。然后利用DNA重组技术依次亚克隆入改造过的真核表达载体pcD-NA3(切除CMV启动子),构建成牛乳腺特异表达载体。获得的重组载体经限制性内切酶酶切鉴定,测序验证等表明,成功构建了牛乳腺特异表达载体。
The high-fidelity PCR method had been used to amplificate 1.8 and 1.1 kb 5＇and 3＇ regulatory sequences of bovine casein gene in mammary grand. Then it was cloned into TA vector. Veritficated by the restriction enzyme digestion and PCR method,after sequence on NCBI with blast, the results indicated that the tragments have the homology of 97.0% and 99.0% respectively with the correspondingly region of boving beta-casein. It is diseribed that the restructuring carrier had succeeded in cloning the area of control casein gene 5＇and 3＇, the recombinant DNA technology had been used to subclone into the modified eukaryotic expression vector pcDNA3 （removal of CMV promoter）, verificated by the restriction enzyme digestion and sequence analysis, the mammary grand specific expression vector of cattle had been constructed.
出处
《中国畜牧兽医》
CAS
北大核心
2009年第8期54-56,共3页
China Animal Husbandry & Veterinary Medicine
基金
贵州省优秀科技教育人才省长专项基金资助项目。黔科教办[2007]03号
