重组大肠杆菌高效分泌表达α-1-6-葡聚糖酶
Construction of a Recombinant Escherichia Coli DH5α for Dextranase Expression
Abstract
以Plf3为模板,PCr扩增得到葡聚糖酶的启动子,并将启动子基因重新连接到切除启动子基因及葡聚糖酶基因的Plf3载体上,构建Plf3-PrO载体.采用PCr的方法扩增朱黄青霉总dnA中的α-1-6-葡聚糖酶基因,并插入到Plf3-PrO载体上,构建Plf3-PrO-dEX质粒,以大肠杆菌dH5α为宿主菌,酶切验证及PCr验证均表明成功构建了Plf3-PrO-dEX质粒,且重组质粒中的α-1-6-葡聚糖酶基因与朱黄青霉中的α-1-6-葡聚糖酶基因有93%的同源性. The promoter of dextranase was PCR-amplified from pLF3 vector and inserted into pLF3 plasmid,from which the genes encoding a promoter and β-glucanase had been removed,yielding PLF3-Pro.The DNA encoding the dextranase was PCR-amplified from the total DNA of Penicillium minioluteum MUCL 38929 and cloned into pLF3-Pro vector.The recombinant plasmid pLF3-Pro-Dex was constructed and checked through PCR,restriction enzyme digestion and DNA sequencing.Results shows that the recombinant plasmid pLF3-Pro-Dex is successfully constructed.