摘要
【目的】构建一个产朊假丝酵母(Candida utilis,C.utilis)整合表达载体。【方法】该载体以质粒pBR322为骨架,包括3-磷酸甘油醛脱氢酶(GAP)启动子和终止子、放线菌酮(CYH)抗性基因和18SrDNA介导的同源整合区。再以木聚糖酶基因为目标基因,插入pGLR9K载体上,构建重组表达载体,电击转化C.utilis。对阳性转化子进行酶活测定,检测其表达情况。【结果】转化子的胞内外都可检测到木聚糖酶酶活,酶活可达60U/mL。【结论】本实验构建了一个C.utilis载体,并用此载体表达了木聚糖酶基因,本研究将为产朊假丝酵母工程菌在饲料添加剂及食品行业中的应用提供又一个新的实验平台。
[Objective]Xylanase is ahemicellulase. It can reduce xylan, which is the abundance resource on our earth. It is important in industries to obtain recombinant strains secreting xylanase to degrade bemicelluloses and produce xylose as desired products. We constructed an integrative vector of Candia utilis for xylanase expression.[Metbods]On the basis of plasmid pBR322, an integrative expression vector pGLR9K for Candida utilis was constructed that contained strong promoter of GAP, 18S rDNA sequence for homologous recombination and mutated L41 gene as CYH resistant marker. Recombinant expression vector pGLXR was constructed by adding xylanase gene to the pGLR9K and then was transferred into C. utilis.[Results]Both intracellular and extracellular xylanase activities were detected in transgenic strains.[Conclusion] This system can be used for exogenous genes expression in transgenic C. utilis.
出处
《微生物学报》
CAS
CSCD
北大核心
2009年第3期316-323,共8页
Acta Microbiologica Sinica
基金
国家“973项目”(2004CB719704)
新疆自治区高技术研究与发展计划项目(200511107)
国家农业科技成果转化资金项目(2006GB2G400336)~~
关键词
产朊假丝酵母
同源整合
载体构建
整合表达
木聚糖酶
Candida utilis
Homologous recombination
Integrative expression
Vector construction
Xylanase.
