毕赤酵母直接基因敲除方法的研究

Recent Advances in Researches on Pichia pastoris directly knockout

甲醇营养型巴斯德毕赤酵母已被广泛应用于外源蛋白的表达,但是作为一个外源蛋白表达系统,其还有待进一步完善。进一步提高外源蛋白的表达量以及形成折叠正确的目的蛋白,成为了改造毕赤酵母菌株的一个热点。基因缺失,是一个常用的改造和构建新的毕赤酵母重组菌株常用的方法。巴斯德毕赤酵母直接基因缺失方法主要分为两类:有标记的插入替换和无标记的Pop-In/Pop-Out。本文系统地介绍了这两种方法,分别比较了各自的优缺点,为它们在巴斯德毕赤酵母中进行基因敲除提供了一定参考意义。

The MethylotrophicPichia pastorishas been widely used in the expression of heterologous proteins, but it has yet to be further improved. Researches of enhancing the expression of heterologous proteins and the correctly folding mechanism of specific protein have gained much attention during last several years. Gene disruption is a commonly used technique to reconstruct a newPichia pastoris- strain, and there are normally two ways to achieve this： gene replacement with marker and markerfree pop-in/pop-out. Here the paper summarizes these two methods, compares pros and cons of each of them, and provides reference for other researchers.