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通过在pGLR9K中进行18S rDNA靶向同源重组,木聚糖酶基因在产朊假丝酵母中的有效表达。

The effective expression of xylanase gene in Candida utilis by 18S rDNA targeted homologous recombination in pGLR9K.

作者信息

Wei Wang, Hong-Lan Yang, HuiFang Bao, Daoyuan Zhang, Qi-mu-ge Shan, Woof Andrew J

机构信息

Insititute of Microbiology of Xinjiang Academy of Agricultural Sciences, 830091, Urumqi, China.

出版信息

Mol Biol Rep. 2010 Jul;37(6):2615-20. doi: 10.1007/s11033-009-9786-x.

Abstract

In order to test whether 18S rDNA can influence positively xylanase gene effective expression in the yeast of Candida utilis, a targeting vector pGLR9K-XA was constructed by adding an interested gene xynA from Streptomyces olivaceoviridis into the vector pGLR9K which is constructed by ourselves. pGLR9K contains the 18S rDNA, GAP promoter and CYH resistance gene sequence, all of which is from C. utilis. Then the vector pGLR9K-XA was transformed into C. utilis. To test the vector and transformed system, PCR, Southern blot and DNS methods were used. The results showed that xylanase gene can be detected in the chromosome DNA of recombinant C. utilis and the enzyme activity of xylanase is up to 60 IU ml(-1) in the study. It is suggested that this system can be used to express exogenous genes in C. utilis as a bioreactors. This is the first report that xylanase gene was expressed in C. utilis.

摘要

为了检测18S rDNA是否能正向影响产朊假丝酵母中木聚糖酶基因的有效表达,通过将来自橄榄绿链霉菌的目的基因xynA添加到我们自己构建的载体pGLR9K中,构建了靶向载体pGLR9K-XA。pGLR9K包含18S rDNA、GAP启动子和CYH抗性基因序列,所有这些均来自产朊假丝酵母。然后将载体pGLR9K-XA转化到产朊假丝酵母中。为了检测该载体和转化系统,使用了PCR、Southern印迹和DNS方法。结果表明,在重组产朊假丝酵母的染色体DNA中可检测到木聚糖酶基因,并且在该研究中木聚糖酶的酶活性高达60 IU ml(-1)。表明该系统可作为生物反应器用于在产朊假丝酵母中表达外源基因。这是关于木聚糖酶基因在产朊假丝酵母中表达的首次报道。

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