基于重叠延伸PCR构建幽门螺杆菌MreB重组载体
发布时间：2019-04-18 14:58
【摘要】：[目的]基于重叠延伸PCR技术,构建串联亲和层析标签(TAP)标记的幽门螺杆菌骨架蛋白Mre B的重组质粒。[方法]将幽门螺杆菌Mre B基因终止密码子TAA前DNA序列(Mre Ba)、TAP和终止密码子TAA后的DNA序列(Mre Bb),通过重叠延伸PCR进行连接,形成大小约3.1 kb的融合片段Mre BCF;Mre BCF片段经XhoⅠ酶切、纯化后,克隆到经SmaⅠ和XhoⅠ双酶切的线性载体p K18mob Sac B上。[结果]PCR、酶切及DNA测序的结果表明,重组质粒p K18Mre BCF包含大小约为740 bp、1 400 bp和1 000 bp的三个片段(Mre Ba、TAP和Mre Bb),并且这三个片段的接头连接及核苷酸序列完全正确。[结论]利用重叠延伸PCR可对多个片段进行无缝连接,简便、高效地构建重组质粒;成功构建了重组质粒p K18Mre BCF,为将来幽门螺杆菌Mre B蛋白功能复合体的分离和鉴定奠定了基础。
[Abstract]:[objective] to construct the recombinant plasmid of Mre B, a cytoskeleton protein of Helicobacter pylori labeled by tandem affinity chromatography (TAP), based on the overlap extension PCR technique. [methods] the pre-TAA DNA sequence (Mre Ba), TAP of Helicobacter pylori Mre B gene and the DNA sequence (Mre Bb), after terminating codon TAA were connected by overlapping extended PCR to form a fusion fragment Mre BCF; of about 3.1 kb in size. The Mre BCF fragment was digested by Xho 鈪，
本文编号：2460126
[Abstract]:[objective] to construct the recombinant plasmid of Mre B, a cytoskeleton protein of Helicobacter pylori labeled by tandem affinity chromatography (TAP), based on the overlap extension PCR technique. [methods] the pre-TAA DNA sequence (Mre Ba), TAP of Helicobacter pylori Mre B gene and the DNA sequence (Mre Bb), after terminating codon TAA were connected by overlapping extended PCR to form a fusion fragment Mre BCF; of about 3.1 kb in size. The Mre BCF fragment was digested by Xho 鈪，
本文编号：2460126
本文链接：https://www.wllwen.com/xiyixuelunwen/2460126.html
最近更新
教材专著
