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α-乙酰乳酸脱羧酶在啤酒生产中能加快啤酒成熟,有重要的应用价值。本研究将枯草芽孢杆菌启动子P43克隆到质粒pUC19-ALDC中的α-乙酰乳酸脱羧酶基因之前,得到重组质粒pUC19-P43-ALDC。重组质粒pUC19-P43-ALDC与质粒pMLK83-BN同源重组,筛选得到枯草芽孢杆菌整合质粒pMLK83-ALDC。用此整合质粒转化枯草芽孢杆菌1A751,挑选出新霉素抗性且无淀粉酶活性的重组菌株。此菌株用LB培养基在37℃、220r/min摇瓶培养过夜,测得α-乙酰乳酸脱羧酶活力为15.6U/mL,说明整合的α-乙酰乳酸脱羧酶基因能够在重组菌株中稳定传代和表达。本研究首次在枯草芽孢杆菌中用整合型的方式重组表达了α-乙酰乳酸脱羧酶,提出了一种有潜力的生产α-乙酰乳酸脱羧酶的新方法。
Abstract：α-acetolactate decarboxylase plays an important applied role in speeding up the maturation of beer in beer production. In this study we obtained the recombinant plasmid pUC19-P43-ALDC by cloning promoter P43 of Bacillus subtilis in the front of α-acetolactate decarboxylase gene in plasmid pUC19-ALDC, which was ho-mologous recombinated with plasmid pMLK83-BN and obtaining the integration plasmid pMLK83-ALDC of Bacillus subtilis. Then we transformed the integration plasmid into Bacillus subtilis 1A751, and selected the re-combinant strain by neomycin resistance screening and amylase activity negative screening. After overnight flask-ing at 37℃, 220 r/min in LB medium, the α-acetolactate decarboxylase activities was 15.6 U/mL with this strain. The integrated α-acetolactate decarboxylase gene would be passaged and expressed stably in the recombinant strain. In this study, we expressed recombinant α-acetolactate decarboxylase gene integratedly in Bacillus subtilis for the first time, and proposed a potentially new way of producing α-acetolactate decarboxylase.
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基本信息:
中图分类号:Q78
引用信息:
[1]廖东庆,陈发忠,岳田芳,等.α-乙酰乳酸脱羧酶基因在枯草芽孢杆菌中的整合型表达[J].基因组学与应用生物学,2010,29(05):843-848.
基金信息:
广西科学研究与技术开发计划项目(桂科产10100021-2)资助
2010-10-28
2010-10-28