绵羊β-乳球蛋白基因5′端及上游调控序列的PCR扩增的方法学研究  

Study on Amplification of BLG 5′-Flanking and Regulatory Sequence by PCR

作　　者：李雪玲[1] 郭继彤[1] 于海泉[1] 王达珍[1] 张肇英[1] 

机构地区：[1]内蒙古大学生命科学学院,内蒙古呼和浩特010021

出　　处：《内蒙古大学学报（自然科学版）》2001年第1期79-82,共4页Journal of Inner Mongolia University：Natural Science Edition

基　　金：国家自然科学基金资助项目，批准号39360034

摘　　要：本文对绵羊 β-乳球蛋白基因 5′端及上游调控序列的 PCR方法扩增进行了研究 .经比较分析 ,将从羊血中提取的绵羊基因组 DNA的模板用量定为 4μl( 0 .4μg/μl) ,Taq DNA聚合酶用量为 0 .83μl( 3u/μl) ,变性为 94℃ 1 min,退火为 64℃ 1 m in,72℃延伸 2 min,循环次数为32次 ,获得的 PCR产物经电泳检测 ,条带明亮 ,特异性高 ,大小为 898bp.经序列分析发现 ,与已知基因组序列一致性达 99%以上 ,可用于指导外源基因在转基因动物乳腺中表达 .This study was carried on the amplification of BLG 5 ′ flanking and regulatory sequence by PCR. The template was the sheep genomic DNA extracted from blood of sheep. The denaturation temperature was 94 ℃ 1 min,the annealling temperature was 64℃ 1min,the extention temperature was 72 ℃ 2 mins. After 32 cycles,the product was analyzed by agarose gel electrophoresis,and sequence assay ,and extracted and used to construct new gene with hCD14. The results shows that this PCR product shares 99% nucleotide sequence with the reported nucleotide sequence of the genomic ovine BLG and can direct the mammary specific expression of hCD14 in transgenic mice.

关 键 词：绵羊β-乳球蛋白基因(BLG)5′端及上游调控序列 PCR 

分 类 号：S826.2[农业科学—畜牧学] Q78[农业科学—畜牧兽医]