Dataset Information

Marker-free genome editing in Ustilago trichophora with the CRISPR-Cas9 technology.

ABSTRACT: In this communication, we report the adaptation of the CRISPR-Cas9 technology in Ustilago trichophora prototrophic wild-type isolate obtained from its natural host Echinochloa crus-galli. The established CRISPR vector and method enable a rapid and marker-free introduction of Cas9-induced non-homologous end-joining (NHEJ) dependent mutation at the targeted gene. Moreover, the method allows a specific modification of the chromosomal DNA sequence by Cas9-induced homologous recombination using short DNA repair templates. The results demonstrate the applicability of the CRISPR-Cas9 technology in U. trichophora for both gene knock-out by the NHEJ pathway and specific gene modification by templated genome editing, paving the way for rapid metabolic engineering of this Ustilago species for industrial applications.

SUBMITTER: Huck S

PROVIDER: S-EPMC6546355 | biostudies-literature | 2019 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Marker-free genome editing in Ustilago trichophora with the CRISPR-Cas9 technology.

Huck Simon S Bock Josephine J Girardello Jörg J Gauert Marc M Pul Ümit Ü

RNA biology 20180810 4

In this communication, we report the adaptation of the CRISPR-Cas9 technology in Ustilago trichophora prototrophic wild-type isolate obtained from its natural host Echinochloa crus-galli. The established CRISPR vector and method enable a rapid and marker-free introduction of Cas9-induced non-homologous end-joining (NHEJ) dependent mutation at the targeted gene. Moreover, the method allows a specific modification of the chromosomal DNA sequence by Cas9-induced homologous recombination using short ...[more]

PMID: 29996713

Similar Datasets

Project description:The discovery and exploitation of the prokaryotic adaptive immunity system based on clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (Cas) proteins have revolutionized genetic engineering. CRISPR-Cas tools have enabled extensive genome editing as well as efficient modulation of the transcriptional program in a multitude of organisms. Progress in the development of genetic engineering tools for the genus Clostridium has lagged behind that of many other prokaryotes, presenting the CRISPR-Cas technology an opportunity to resolve a long-existing issue. Here, we applied the Streptococcus pyogenes type II CRISPR-Cas9 (SpCRISPR-Cas9) system for genome editing in Clostridium acetobutylicum DSM792. We further explored the utility of the SpCRISPR-Cas9 machinery for gene-specific transcriptional repression. For proof-of-concept demonstration, a plasmid-encoded fluorescent protein gene was used for transcriptional repression in C. acetobutylicum Subsequently, we targeted the carbon catabolite repression (CCR) system of C. acetobutylicum through transcriptional repression of the hprK gene encoding HPr kinase/phosphorylase, leading to the coutilization of glucose and xylose, which are two abundant carbon sources from lignocellulosic feedstocks. Similar approaches based on SpCRISPR-Cas9 for genome editing and transcriptional repression were also demonstrated in Clostridium pasteurianum ATCC 6013. As such, this work lays a foundation for the derivation of clostridial strains for industrial purposes.ImportanceAfter recognizing the industrial potential of Clostridium for decades, methods for the genetic manipulation of these anaerobic bacteria are still underdeveloped. This study reports the implementation of CRISPR-Cas technology for genome editing and transcriptional regulation in Clostridium acetobutylicum, which is arguably the most common industrial clostridial strain. The developed genetic tools enable simpler, more reliable, and more extensive derivation of C. acetobutylicum mutant strains for industrial purposes. Similar approaches were also demonstrated in Clostridium pasteurianum, another clostridial strain that is capable of utilizing glycerol as the carbon source for butanol fermentation, and therefore can be arguably applied in other clostridial strains.

Dataset Information

Marker-free genome editing in Ustilago trichophora with the CRISPR-Cas9 technology.

Publications

Marker-free genome editing in Ustilago trichophora with the CRISPR-Cas9 technology.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets