CRISPR/Cas9-RNA interference system for combinatorial metabolic engineering of Saccharomyces cerevisiae.
Yeast
; 36(5): 237-247, 2019 05.
Article
em En
| MEDLINE
| ID: mdl-30953378
The yeast Saccharomyces cerevisiae is widely used in industrial biotechnology for the production of fuels, chemicals, food ingredients, food and beverages, and pharmaceuticals. To obtain high-performing strains for such bioprocesses, it is often necessary to test tens or even hundreds of metabolic engineering targets, preferably in combinations, to account for synergistic and antagonistic effects. Here, we present a method that allows simultaneous perturbation of multiple selected genetic targets by combining the advantage of CRISPR/Cas9, in vivo recombination, USER assembly and RNA interference. CRISPR/Cas9 introduces a double-strand break in a specific genomic region, where multiexpression constructs combined with the knockdown constructs are simultaneously integrated by homologous recombination. We show the applicability of the method by improving cis,cis-muconic acid production in S. cerevisiae through simultaneous manipulation of several metabolic engineering targets. The method can accelerate metabolic engineering efforts for the construction of future cell factories.
Palavras-chave
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Saccharomyces cerevisiae
/
Interferência de RNA
/
Engenharia Metabólica
/
Sistemas CRISPR-Cas
Idioma:
En
Ano de publicação:
2019
Tipo de documento:
Article