PCR & Go: A Pre-installed Expression Chassis for Facile Integration of Multi-Gene Biosynthetic Pathways.
Front Bioeng Biotechnol
; 8: 613771, 2020.
Article
em En
| MEDLINE
| ID: mdl-33520963
ABSTRACT
The introduction of multi-gene metabolic pathways is generally the first step for the construction of microbial cell factories and plays an essential role in metabolic engineering and synthetic biology. Here, we developed a "PCR & Go" system for facile integration and assembly of multi-gene pathways into the chromosome of Saccharomyces cerevisiae. The core component of the "PCR & Go" system was an expression chassis, where eight promoter/terminator pairs were pre-installed into the yeast chromosome and PCR amplified gene fragments could be inserted directly for functional expression. In combination with the CRISPR/Cas9 system and a gRNA plasmid library, the ß-carotene (three genes), zeaxanthin (four genes), and astaxanthin (five genes) biosynthetic pathways were integrated and assembled into the yeast genome with an efficiency of ~93, ~85, and 69%, respectively, using PCR amplified gene fragments with ~40 bp homology arms in a single step. Therefore, the "PCR & Go" system can be used for fast construction of yeast cell factories harboring multi-gene pathways with high efficiency and flexibility.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Idioma:
En
Revista:
Front Bioeng Biotechnol
Ano de publicação:
2020
Tipo de documento:
Article
País de afiliação:
China
País de publicação:
CH
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SUIZA
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SUÍÇA
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SWITZERLAND