Construction of a herbal plant cDNA library for functional expression in Saccharomyces cerevisiae

• Main Authors: Ren-zong Jheng, 鄭仁棕
• Other Authors: none
• Format: Others
• Language: zh-TW
• Published: 2012
• Online Access: http://ndltd.ncl.edu.tw/handle/73435157841656588980

碩士 === 慈濟大學 === 生命科學系碩士班 === 100 === As the first step towards the production of express of triterpene cyclase in yeast, we have constructed a PCR-amplified cDNA library from Artemisia argyi, and we tried five different expression systems to express the cDNA more effectively. Three of the cloning systems were modified from the Gateway technology and the other two were converted from the TA cloning technique which create expression clones without using restriction enzymes. For the modified gateway technology, linear cDNAs were designed to directly ligate with vectors through the lambda LR or BP recombination. By using the TA cloning techniques, blunt-end attachment with dTTP and restriction enzyme AhdI were used to create cloning T-vectors for direct ligation with PCR-amplified plant cDNAs. Though these cloning systems were constructed , the cloning efficiency is not good enough yet to express a potent plant cDNA library for the cloning of triterpene cyclase. The future work will be focused on the improvement of the cloning efficiency for cDNA library construction. This expression system is universal for the plant, especially from traditional Chinese herbal plants without detailed genetic or molecular information.