CRISPR/Cas9‐RNA interference system for combinatorial metabolic engineering of Saccharomyces cerevisiae. Issue 5 (13th June 2019)

• Record Type: Journal Article
• Authors: Kildegaard, Kanchana Rueksomtawin; Tramontin, Larissa Ribeiro Ramos; Chekina, Ksenia; Li, Mingji; Goedecke, Tobias Justus; Kristensen, Mette; Borodina, Irina
• Other Names: Sibirny Andriy guestEditor.
• Is Part Of: Yeast. Volume 36:Issue 5(2019:May)
• Page Start: 237
• Page End: 247
• Publication Date: 2019-06-13
• Subjects: cis, cis‐muconic acid -- CRISPR/Cas9 -- genome editing -- metabolic engineering -- RNA interference -- USER (uracil‐specific excision reagent) -- Saccharomyces cerevisiae; Yeast -- Periodicals; Yeasts -- Periodicals; Yeasts -- genetics -- Periodicals; Electronic journals; 547
• DOI: 10.1002/yea.3390
• Languages: English
• ISSNs: 0749-503X

Abstract

Abstract: 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.