雨生红球藻cDNA表达文库的构建与初步分析

Construction and Identification of cDNA Expression Library from Haematococcus pluvialis

为了筛选雨生红球藻虾青素合成过程中的关键酶基因的反式调节因子基因,以雨生红球藻的绿细胞为材料,提取了高质量的总RNA,并分离纯化了mRNA,经过反转录后的得到cDNA,构建了以λ-ZAPExpress为载体的表达型cDNA文库。经检测,所构建的文库的滴度为5.1×105,重组率为100%。用PCR方法对文库的质量进行了鉴定,文库的平均插入片断的大小为1.7kb。雨生红球藻cDNA表达文库的构建为研究虾青素的代谢工程奠定了基础。

To select the genes of trans-factors involved in the regulation of the synthesis of astaxanthin, the total RNA was extracted from H.pluvlialis, the purified mRNA was reversed to cDNA, and a cDNA expression library was constructed using λ ZAP Express vector from motile cells of Haernatococcus pluvlialis. The test result showed tl^e titer of the cDNA expression library was 5.1×10^5 pfu/ml and the efficiency of recombination was 100%. Using PCR to test the quality of the constructed library, the results showed that the average size of the cDNA insert fragments was 1.7 kb. The constructed cDNA Expression library from H. pluvialis has laid foundation for the research of metabolic engineering of astaxanthin.