期刊
YEAST
卷 31, 期 2, 页码 61-66出版社
WILEY
DOI: 10.1002/yea.2994
关键词
Schizosaccharomyces pombe; fission yeast; protoplasts; spheroplasts; Lallzyme; zymolyase
资金
- Spanish Ministerio de Ciencia e Inovacion [BFU2010-21310]
- Junta de Andalucia (Proyecto de Excelencia) [P09-CTS-4697]
- Spanish Ministerio de Educacion (Juan de la Cierva Programme)
Fungal cells including yeasts are surrounded by cell wall that counteracts turgor pressure and prevents cell lysis. Many yeast experiments, including genetic manipulation of sterile strains, morphogenesis studies, nucleic acid isolation and many others, require mechanical breakage or enzymatic removal of the cell wall. Some of these experiments require the generation of live cells lacking cell walls, called protoplasts, that can be maintained in osmostabilized medium. Enzymatic digestion of cell wall proteoglycans is a commonly used method of protoplast preparation. Currently existing protocols for fission yeast cell wall digestion are time consuming and not very efficient. We developed a new rapid method for fission yeast protoplast preparation that relies on digesting cell walls with Lallzyme MMX commercial enzyme mix, which produces protoplasts from all cells in less than 10min. We demonstrate that these protoplasts can be utilized in three commonly used fission yeast protocols. Thus, we provide the fission yeast community with a robust and efficient plasmid extraction method, a new protocol for diploid generation and an assay for protoplast recovery that should be useful for studies of morphogenesis. Our method is potentially applicable to other yeasts and fungi. Copyright (c) 2013 John Wiley & Sons, Ltd.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据