Endogenous 2μ Plasmid Editing for Pathway Engineering in Saccharomyces cerevisiae.

ABSTRACT: In Saccharomyces cerevisiae, conventional 2μ-plasmid based plasmid (pC2μ, such as pRS425) have been widely adopted in pathway engineering for multi-copy overexpression of key genes. However, the loss of partition and copy number control elements of yeast endogenous 2μ plasmid (pE2μ) brings the issues concerning plasmid stability and copy number of pC2μ, especially in long-term fermentation. In this study, we developed a method based on CRISPR/Cas9 to edit pE2μ and built the pE2μ multi-copy system by insertion of the target DNA element and elimination of the original pE2μ plasmid. The resulting plasmid pE2μRAF1 and pE2μREP2 demonstrated higher copy number and slower loss rate than a pC2μ control plasmid pRS425RK, when carrying the same target gene. Then, moving the essential gene

SUBMITTER: Zeng BX 

PROVIDER: S-EPMC7921170 | biostudies-literature | 2021

REPOSITORIES: biostudies-literature