树干毕赤酵母基因组文库的构建及纤维二糖酶基因的筛选

Construction of Pichia stipitis Genomic Library and Cellobiase Gene Screening

摘要:

首次构建了树干毕赤酵母DNA基因组文库,该文库包含3 000个克隆,插入片段长度为0.5～8.0 kb。树干毕赤酵母染色体基因为15 441 179 bp,该文库覆盖1.08倍树干毕赤酵母染色体基因,筛选出任一基因或序列的概率为97%。提取文库中的质粒醋酸锂法转化酿酒酵母W303-1A,涂布于纤维二糖为唯一碳源的平板,通过纤维二糖酶对底物纤维二糖的底物特异性筛选文库,首次获得树干毕赤酵母来源的一个1.82 kb的纤维二糖酶基因。

Abstract:

In this study,a genomic library for Pichia stipitis was firstly constructed.The library covered 1.08 times of the15 441 179bp genomic DNA of the strain,and contained 3 000 clones with the insertion sizes ranging from 0.5 to 8.0 Kb.It has a propability of 97% for screening a random gene form this library.The library plasmids were extracted and transformed into Saccharomyces cerevisiae W303-1A by LiAc method.At last,a 1.82 Kb cellobiase gene of P.stipitis was firstly characterized by means of screening the transformants on cellobiose plates.