【摘要】 从菊粉酶酶源菌株克鲁维酵母GKluyveromycesmarxianus)Y109中提取菊粉酶基因,PCR扩增Y109的菊粉酶基因inuB1,核苷酸序列测定分析表明inuB1的阅读框架全长1665bp,没有内含子,其中存在4个潜在的N 糖基化位点。以pPIC9K为表达载体,构建inuB1的酵母转化载体,转化毕赤酵母(Pichiapastoris)GS115,获得菊粉酶基因工程菌GS115/inuB1。GS115/inuB1甲醇诱导发酵表达菊粉酶,inuB1基因重组胞外菊粉酶表观分子量为90kD(SDS PAGE),比酶活性是12.29U/mg。更多还原

【Abstract】 We isolate Inulinase gene inuB1 from Kluyveromyces marxianus Y109 and integrate it into the polycloning site expressed in vector pPIC9K. Experiment results reveal that the Open Reading Frames (ORFs) of inuB1 is 1 665?bp. We find no intron and four potential sites for Nlinked glycosylation in the sequence. The pPIC9K/inuB1 construct is used to transform Pichia pastoris GS115 (His-, Mut+). The gene inuB1 is integrated into the genomic DNA of GS115 by insertion into a single site for recombination, yielding the recombinant GS115/inuB1. Inulinase is induced by methanol.Its apparent molecular weight is 90kDa (SDSPAGE) and specific activity approaches 12.29?U/mg.更多还原