Department of Biological Sciences, University of the Pacific, Stockton, CA, USA.
Methods Mol Biol. 2022;2513:113-120. doi: 10.1007/978-1-0716-2399-2_7.
During the past three decades, the methylotrophic yeast Pichia pastoris (recently reclassified as Komagataella phaffii) has gained widespread acceptance as a system of choice for heterologous protein expression. One of the reasons that this yeast is used so frequently is the simplicity of techniques required for its molecular genetic manipulation. There are several different protocols available for introducing DNA into P. pastoris using electroporation or heat shock. We describe here a shortened protocol for cell preparation and transformation that works reliably with either prototrophic markers or antibiotic selection in this host. This procedure utilizes the most efficient portions of the electroporation and heat-shock transformation protocols to yield a method that is both time-saving and effective.
在过去的三十年中，甲醇营养型酵母巴斯德毕赤酵母（最近重新归类为巴斯德毕赤酵母）已被广泛认可为用于异源蛋白表达的首选系统。这种酵母如此频繁地被使用的原因之一是其分子遗传操作所需的技术非常简单。有几种不同的方案可用于用电穿孔或热休克将 DNA 导入巴斯德毕赤酵母。我们在这里描述了一种简化的细胞制备和转化方案，该方案可与该宿主中的营养缺陷型标记或抗生素选择可靠地配合使用。该方法利用了电穿孔和热休克转化方案中最有效的部分，从而产生了一种既省时又有效的方法。
