摘要
目的构建卡介苗菌株Hsp16.3基因打靶载体。方法体外培养卡介苗菌株并提取基因组DNA,扩增Hsp16.3基因两侧序列5'-Hsp16.3和3'-Hsp16.3,分别插入到pKO质粒载体预定位点中构建Hsp16.3基因置换型打靶载体,筛选并进行PCR、双酶切鉴定及测序鉴定。结果构建的卡介苗菌株Hsp16.3基因打靶载体,经PCR、双酶切鉴定及测序证实pKO质粒中的2个插入片段为所需目的基因。结论成功构建出卡介苗菌株Hsp16.3基因打靶载体。
【Objective】 To construct a targeting vector for the gene encoding Hsp16.3 of Mycobacterium BCG.【Methods】 Mycobacterium BCG was cultured in vitro and the genome DNA was extracted.In order to obtain a gene targeting vector pKO-Hsp16.3,two fragments（5＇-Hsp16.3,3＇-Hsp16.3） flanking the Hsp16.3 region were amplified by PCR and inserted into pKO plasmid.The recombinant pKO-Hsp16.3 plasmid was further identified by polymerase chain reaction（PCR）,digested by double restriction enzymes and sequencing.【Results】 The targeting vector we constructed was identified with PCR,restriction enzyme digestion and sequencing,and the two gene fragments that have been inserted into the pKO plasmid were the target genes.【Conclusions】 A targeting vector for the gene encoding Hsp16.3 of Mycobacterium BCG was constructed successfully.
出处
《中国现代医学杂志》
CAS
CSCD
北大核心
2012年第11期8-13,共6页
China Journal of Modern Medicine
基金
国家自然科学基金(No:30960355)
石河子大学科学技术研究发展计划"自然科学与计划创新"重点项目(No:ZRKX2010ZD01)
