灭蚊真菌贵阳腐霉基因组文库的构建

Construction of a Genomic Library of Pythium guiyangense Su,A mosquito-killing Fungus

目的:建立贵阳腐霉(Pythium guiyangense Su)基因组文库以便克隆该真菌的基因特别是与灭蚊毒力有关的基因。方法:用限制性内切酶Sau3AI消化贵阳腐霉基因组DNA,同时用限制性内切酶BamHI消化质粒pBluescript SK(-),再将其去磷酸化;将基因组的酶切产物连接到载体上,再转化大肠杆菌DH5α。结果:所建贵阳腐霉的基因组文库共包含11 520个克隆。经蓝白斑筛选、PCR验证以及双酶切验证,文库平均插入片段长度为1.2 kb,覆盖率约为贵阳腐霉基因组的3倍。结论:建立了贵阳腐霉的基因组文库,可以从中克隆与贵阳腐霉灭蚊毒力有关的基因。

Objective：To construct a genomic library of Pythium guiyangense Su,strain 1938 GY.Methods：The digested genomic DNA with Sau3AI was inserted into pBluescript SK（-） plasmid which was digested with BamHI and treated with calf intestinal alkaline phosphatase.Next,the recombinant plasmids were transferred into E.coli DH5α.Results：A genomic DNA library was constructed which included 11520 clones with average insert sizes about 1.2 kb.Conlusions：The results of this experiment provide a source for screening genes of P.guiyangense,and lay basis for further work of genetic engineering of this fungus.