SMART技术构建红曲霉全长cDNA文库

Construction of Monascus full-length cDNA library with SMART approach

目的构建红曲霉cDNA文库，为红曲霉功能基因的研究以及筛选、克隆红曲霉次生代谢产物合成途径相关基因奠定基础。方法采用SMART（ switching mechanism at 5 ＇end of RNA transcript）技术构建红曲霉全长cDNA文库，经涂平板和酶切反应测定文库的克隆数、重组率，PCR测定插入片断的大小。结果原始文库的库容为2．03×10^5，重组率达94％。插入片段大小多在0．7—2．0kb，平均大小在1kb左右。结论所构建文库的代表性和重组片段的序列完整性达到了用于目的基因的分离筛选和克隆表达的建库要求。

Objective Construction of Monascus cDNA Library provide foundations for the research of Monascus functional genes and screening, cloning genes related to the synthesis pathway of secondary metabolites in Monascus. Methods A cDNA library was constructed based on SMART system. The number of the clones and the recombination rate of the library were determined by plate coating and restrictive digestion, and the size of inserted fragment was determined by PCR. Results The primary library capacity was 2.03 × 10^5 , average inserted fragment size was about 1 000 bp and the percentage of recombination were 94%. Conclusion The library met the requirement for cloning target genes and expressing target proteins.