摘要
目的克隆Pythium sp.CY1938菌株类枯草菌素蛋白酶Pr1基因。方法在已经克隆得到的Pythi- um sp.GY1938菌株类枯草菌素蛋白酶cDNA片段的基础上,首先通过PCR扩增得到与其相对应的基因组DNA片段,然后分别通过Mini基因组DNA文库法和Panhandle PCR法扩增其上游和下游未知片段,最后通过PCR法扩增得到类枯草菌素蛋白酶基因片段。结果克隆得到1 519bp Pr1蛋白酶基因序列YP1977.DNAassist软件分析其中含有一个969bp的完整的开放阅读框,编码323个氨基酸,NCBI BlastX比对结果显示,该阅读框可能编码的氨基酸序列与多个物种的Pr1蛋白酶都具有较高的同源性。结论YP1977很可能是Pythium sp.CY1938菌株类枯草菌素蛋白酶Pr1基因的部分片段,但仍需要进一步通过构建表达系统加以验证。
Objective To obtain the flail- length genomie DNA sequence of the subtilisin- like protease （Pr1） gene from Pythium sp. GY1938. Methods Basing on the obtained cDNA sequence of the Pr1 from Pythium sp. GY1938, a genomic DNA fragment corresponding to the cDNA was amplified by PCR firstly. Then the upstream and downstream unknown fragments were respectively cloned by mini genomic DNA library and panhandle PCR methods. In the end, the gene fragment of the Pr1 was amplified by PCR. Results A 1 519bp DNA fragment called YP1977, in which there is a 969bp ORF coding 323 amino acids analysed by DNAassist software, of subtilisin - like protease gene from Pythium sp. GY1938 was obtained. Conclusion The fragment, YP1977, is the Pr1 gene from Pythium sp. GY1938 in all probability. However, it must be validated further by the construction of expression system.
出处
《成都医学院学报》
CAS
2006年第1期5-8,共4页
Journal of Chengdu Medical College
