A protocol of rapid laboratory evolution by genome shuffling in Kluyveromyces marxianus

Li Wu; Mengzhu Wang; Genhan Zha; Jungang Zhou; Yao Yu; Hong Lu

doi:10.1016/j.mex.2020.101138

A protocol of rapid laboratory evolution by genome shuffling in Kluyveromyces marxianus

MethodsX. 2020 Nov 12:7:101138. doi: 10.1016/j.mex.2020.101138. eCollection 2020.

Authors

Li Wu^{1

2}, Mengzhu Wang^{1

2}, Genhan Zha^{1

2}, Jungang Zhou^{1

2}, Yao Yu^{1

2}, Hong Lu^{1

2

3}

Affiliations

¹ State Key Laboratory of Genetic Engineering, School of Life Sciences, Fudan University, Shanghai 200438, China.
² Shanghai Engineering Research Center of Industrial Microorganisms, Shanghai 200438, China.
³ Shanghai Collaborative Innovation Center for Biomanufacturing, East China University of Science and Technology, 130 Meilong Road, Shanghai 200237, China.

Abstract

Genome shuffling is a process to combine advantage traits by the recombination of the entire genome and it has been successfully applied in the laboratory evolution of various industrial microorganisms. However, genome shuffling has not been described in Kluyveromyces marxianus (KM), a promising yeast host for the expression of heterologous proteins. In this protocol, genome shuffling in KM is performed by sexual reproduction and is combined with high-throughput screening to obtain high-yielding strains. Notably, the screening of diploid clones risen from one mating mixture is carried out to improve the effectiveness of evolution. Mating-sporulation-mating cycles are repeated to obtain KM strain with ideal traits. •The method combines genome shuffling with high-throughput to achieve strains displaying high yielding of heterologous proteins.•This method can be applied to the genome shuffling of other species when only a few starting strains are available for sexual reproduction.

Keywords: Genome shuffling; Ideal phenotypes; Mating.