牛β-酪蛋白基因的克隆及乳腺特异性表达载体的构建  

Cloning of Bovine Beta-Casein Gene and Construction of Mammary Gland-Specific Expression Vector

作　　者：杨瑞锋[1] 刘金龙[1] 安志兴[1] 李志艳[1] 张涌[1] 

机构地区：[1]西北农林科技大学生物工程研究所,杨凌712100

出　　处：《中国生物工程杂志》2005年第B04期260-263,共4页China Biotechnology

基　　金：国家"863"计划资助项目(2001AA213081)

摘　　要：利用PCR技术分3段克隆了长约9．7kb的牛β-酪蛋白基因,分另4克隆在pGEM-T easy Vector的T位点。经NCBI Blastn分析,与牛β-酪蛋白基因相应区段的同源性为98％。这3段序 列包含完整的5’侧翼序列、前8个外显子及第8个内含子的部分序列。以此构建了两个乳腺特 异性表达栽体:利用第1段和第2段共有的酶切位点将两段融合,作为5’调控区(6．8kb),第3段 和实验室已有的600 bp的加尾信号通过重组PCR拼接作为3’调控区(3．4kb)构建了一乳腺特异 性表达栽体;以第一段作为5’调控区,实验室已有的600bp加尾信号作为3’为调控区构建了另一 乳腺特异性表达载体。可以应用于进一步乳腺生物反应器的研制。Three fragments of bovine beta-casein （about 9.7kb） were cloned by PCR, and then were subcloned into pGEM T easy Vector. After sequence analysis on NCBI with Blastn, results indicated that the fragments have the homology of 98.0% respectively with the correspondingly region of bovine beta-casein. And these three fragments includes integrated 5＇flanking sequence, eight exons and partial the eighth introno Based on them, two mammary gland-specific expression vectors were constructed. One of them use fused fragments of CN1 and CN2 as 5＇promoter, splicing fragments of CN3 and 600bp poly A by recombinant PCR as 3＇promoter. The other used CN1 as 5＇promoter, 600bp poly A as 3＇promoter. Both of them can be applied in preparing mammary gland bioreactor.

关 键 词：牛β-酪蛋白基因 克隆 载体构建 

分 类 号：Q786[生物学—分子生物学] TQ464.7[化学工程—制药化工]