摘要
电击转化由于其高效率而被广泛地应用于酿酒酵母的转化过程中。但是由于菌种退化或是暴露于污染的环境,很多菌种的转化效率会显著地下降2～4个数量级。探索了一种改进的电击转化方法,它借助于单链载体DNA和转化后在YPD培养基中的复苏过程,可以使转化效率比之前曾报道过的已经最优化的用醋酸锂和二硫苏糖醇进行预处理的转化方法效率提高13倍。在毕赤酵母中使用这一改进的方法,可以使转化效率提高高达114倍,在一些已经退化的酿酒酵母菌株中,此方法也可以提高转化效率将近100倍。这一方法将为几乎所有酿酒酵母和毕赤酵母的分子操作提供极大的便利。
Electroporation is widely used for highly efficient transformation of Saccharomyces cerevisiae.As many strains degenerate over time or over environmental exposure,their electroporation efficiencies could drop markedly by 2 to 3 orders of magnitude.Here we describe a modified electroporation procedure which included single stranded carrier DNA for electroporation as well as a post-electroporation growth period in YPD medium,which gave rise to 13 fold enhancement in transformations than an optimized lithium acetate（LiAc） and dithiothreitol（DTT） Pretreatment protocol previously reported.An enhancement of 114 fold on Pichia pastoris transformations has been observed using the modified procedure.The modified method enhanced the transformation efficiencies of some degenerated S.cerevisiae strains by nearly 100 fold.It will enable molecular manipulations in all kinds of S.cerevisiae and P.pastoris strains by allowing propagations of all sorts of DNA libraries made from minute samples.
出处
《中山大学学报（自然科学版）》
CAS
CSCD
北大核心
2010年第3期98-101,共4页
Acta Scientiarum Naturalium Universitatis Sunyatseni
基金
广东省科技计划资助项目(2008B020100001)
广东省自然科学基金资助项目(05003328)
国家自然科学基金资助项目(30370799)
温氏集团和中山大学开放实验室基金资助项目
关键词
电击转化
酿酒酵母
单链载体DNA
转化后复苏
毕赤酵母
electroporation
Saccharomyces cerevisiae
single stranded carrier DNA
post-electroporation growth
Pichia pastoris
