Saccharomyces cerevisiae genetically engineered bacterium for enhancing homologous recombination capability and application thereof

Abstract

The invention discloses a yeast genetically engineered bacterium for enhancing homologous recombination capability and application thereof, and relates to the field of bioengineering. According to the yeast genetically engineered bacterium, yeast is used as an original strain, the yeast genetically engineered bacterium is constructed by introducing genes, and the introduced genes comprise any one of the following groups: (1) an RAD52 gene, an RAD59 gene and an MRE11 gene; (2) an RAD52 gene and an RAD59 gene; (3) an RAD52 gene and an MRE11 gene; (4) an RAD52 gene and an SAE2 gene; (5) an RAD52 gene; and (6) an RAD52 gene, an RAD59 gene, an MRE11 gene and an SAE2 gene. The yeast genetically engineered bacterium provided by the invention can realize simultaneous integration of single, two and three genes of a heterologous gene expression cassette with about 40bp homologous arms, and the efficiencies are respectively up to 100%, about 98% and about 81%.

Description

technical field [0001] The invention relates to the field of bioengineering, in particular to a yeast genetically engineered bacterium with enhanced homologous recombination ability and its application. Background technique [0002] Pichia pastoris can grow in the medium with methanol as the only carbon source and energy source, and has the characteristics of high protein expression. It not only has the strongest methanol-inducible promoter pAOX1 in eukaryotes, but also can carry out high Density fermentation is beneficial to industrial scale production and is considered to be one of the most promising hosts for protein production. Establishment of the methylotrophic yeast Pichia pastoris as a microbial cell factory for the production of fuels, chemicals, and natural products has received increasing attention, especially from methanol as a feedstock. [0003] Considering the stability and metabolic burden of episomal plasmids, genome integration is usually used to efficient...

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Problems solved by technology

However, the number of transformants of KU70 gene knockout strains will be greatly reduced, and there is a possibility of loss of large fragments of chromosome ends (Ito, Y., Watanabe, T., Aikawa, S., Nishi, T., Nishiyama, T., Nakamura, Y., Hasunuma, T., Okubo, Y., Ishii, J., and Kondo, A. Deletion of DNAligase IV homolog confers higher gene targeting efficiency on homologous recombination in Komagataella phaffii. FEMS Yeast Research, 2018, 18. doi: 10.1093 / femsyr / foy074)

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