High-Efficiency Plasmid DNA Transformation in Yeast.
High-Efficiency Plasmid DNA Transformation in Yeast.
复制标题
酵母中的高效质粒 DNA 转化。
DOI：
10.1007/978-1-0716-2399-2_2
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发表时间：
2022
期刊：
影响因子：
--
通讯作者：
Lewis,LKevin
中科院分区：
文献类型：
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作者：
Fitzgerald,O'TaveonR;Rodriguez,NestorD;Lewis,LKevin
将DNA转化为萌芽酵母、酿酒酵母和其他重要工业酵母的细胞，最常用的方法是化学法。目前的方案通常涉及将细胞暴露在含有聚乙二醇(PEG)的溶液中的锂离子中，通常与促进细胞壁和/或细胞膜不稳定的其他试剂一起使用，如二甲基亚砜(DMSO)。最近的工作表明，使用快速且易于扩展的方法，通过早期固定相细胞，即小型过夜液体细胞培养，可以实现高转化效率，以满足高通量项目的需要。在此，我们描述了载体DNA、化学试剂和细胞生长介质，它们允许用质粒或线性DNA片段高效地转化酵母细胞。
Transformation of DNA into cells of the budding yeastSaccharomyces cerevisiaeand other industrially important yeasts is most commonly performed using chemical-based methods. Current protocols typically involve exposure of the cells to lithium ions in a solution containing the crowding agent polyethylene glycol (PEG), often in conjunction with other reagents such as dimethyl sulfoxide (DMSO) that promote destabilization of the cell wall and/or cell envelope. Recent work has demonstrated that it is possible to achieve high transformation efficiencies with early stationary phase cells, i.e., small overnight liquid cell cultures, using methods that are rapid and readily scalable for high-throughput projects. Herein, we describe carrier DNAs, chemical reagents, and cell growth media that permit transformation of yeast cells with either plasmids or linear DNA fragments with high efficiency.