Development of two methods for rapid screening of recombinantPichia pastorisstrains with high-level expression of β-mannanase
来源期刊：中南大学学报(英文版)2015年第11期
论文作者：LI Gu-yue ZHENG Jia ZHOU Hong-bo
文章页码：4162 - 4167
Key words：β-mannanase; Pichia pastoris; high-production strains; hydrolysis hole screening method; micro-plate screening method
Abstract: The yeast Pichia pastoris(P. pastoris) has been used for the expression of heterologous proteins with the significant success. However, it is time-consuming to screen the high expression level of the recombinant P. pastoris directly. Thus, for β-mannanase production, developing the accurate, rapid and inexpensive screening method to substitute random screening is certainly required. A simple method based on the size of hydrolysis hole was described here, but this method was not very accurate that could only be used in preliminary screening. To further improve the accuracy, a micro-plate screening method is established, which appears to be more accurate and effective. The efficiency of this screening method is about 10 times higher than that of the general screening strategy of cultivation in shaking flasks. Two methods presented here can also be used for screening of recombinant Pichia strains with high-level expression of other heterologous protein after modification.
LI Gu-yue(李古月)1, ZHENG Jia(郑甲)1, ZHOU Hong-bo(周洪波)1, 2
(1. School of Minerals Processing and Bioengineering, Central South University, Changsha 410083, China;
2. Key Laboratory of Biometallurgy of Ministry of Education (Central South University), Changsha 410083, China)
Abstract:The yeast Pichia pastoris(P. pastoris) has been used for the expression of heterologous proteins with the significant success. However, it is time-consuming to screen the high expression level of the recombinant P. pastoris directly. Thus, for β-mannanase production, developing the accurate, rapid and inexpensive screening method to substitute random screening is certainly required. A simple method based on the size of hydrolysis hole was described here, but this method was not very accurate that could only be used in preliminary screening. To further improve the accuracy, a micro-plate screening method is established, which appears to be more accurate and effective. The efficiency of this screening method is about 10 times higher than that of the general screening strategy of cultivation in shaking flasks. Two methods presented here can also be used for screening of recombinant Pichia strains with high-level expression of other heterologous protein after modification.
Key words:β-mannanase; Pichia pastoris; high-production strains; hydrolysis hole screening method; micro-plate screening method
