猪源转座子介导的诱变及其在毕赤酵母中的应用。
PiggyBac transposon-mediated mutagenesis and application in yeast Komagataella phaffii.
机构信息
State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, People's Republic of China.
Roche R&D Center (China) Ltd., Shanghai, People's Republic of China.
出版信息
Biotechnol Lett. 2018 Oct;40(9-10):1365-1376. doi: 10.1007/s10529-018-2592-6. Epub 2018 Jul 12.
OBJECTIVE
Around one-fourth of the Komagataella phaffii genes encode hypothetical proteins with unknown functions. However, lack of powerful tools for genetic screening in K. phaffii significantly limits the functional analysis of these unknown genes. Transposon mutagenesis has been utilized as an insertional mutagenesis tool in many other organisms and would be extremely valuable if it could be applied in K. phaffii.
RESULTS
In this study, we investigated in K. phaffii the transposition activity and efficiency of piggyBac (PB) transposon, a DNA transposon from the cabbage looper moth Trichoplusia ni through the integrated-plasmid system. We also designed a binary-plasmid system which could generate stable mutants. Finally we evaluated the quality of this mutagenesis system by a simple screening for functional genes involved in K. phaffii carbon catabolite repression.
CONCLUSIONS
Our results demonstrate that PB-mediated mutagenesis could be a feasible and useful tool for functional gene screening in K. phaffii.
目的
约四分之一的毕赤酵母 Komagataella phaffii 基因编码具有未知功能的假设蛋白。然而，毕赤酵母缺乏强大的遗传筛选工具，这极大地限制了这些未知基因的功能分析。转座子诱变已被用于许多其他生物体中的插入诱变工具，如果它可以应用于毕赤酵母，将会非常有价值。
结果
在这项研究中，我们通过整合质粒系统研究了毕赤酵母中的 piggyBac (PB) 转座子的转座活性和效率，PB 转座子是一种来自甘蓝夜蛾 Trichoplusia ni 的 DNA 转座子。我们还设计了一个可以产生稳定突变体的双质粒系统。最后，我们通过简单筛选参与毕赤酵母碳分解代谢阻遏的功能基因来评估这个诱变系统的质量。
结论
我们的结果表明，PB 介导的诱变可能是毕赤酵母功能基因筛选的一种可行且有用的工具。
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