Cpf1 enables fast and efficient genome editing in Aspergilli

Research output: Contribution to journalJournal article – Annual report year: 2019Researchpeer-review

Documents

DOI

View graph of relations

Background
CRISPR technology has revolutionized fungal genetic engineering by increasing the speed and complexity of the experiments that can be performed. Moreover, the efficiency of the system often allows genetic engineering to be introduced in non-model species. The efficiency of CRISPR gene editing is due to the formation of specific DNA double-strand breaks made by RNA guided nucleases. In filamentous fungi, only Cas9 has so far been used as the CRISPR nuclease. Since, gene editing with Cas9 is limited by its 5′-NGG-3′ protospacer adjacent motif (PAM) sequence, it is important to introduce RNA guided nucleases that depend on other PAM sequences in order to be able to target a larger repertoire of genomic sites. Cpf1 from Lachnospiraceae bacterium employs a PAM sequence composed of 5′-TTTN-3′ and therefore serves as an attractive option towards this goal.

Results
In this study we showed that Lb_cpf1 codon optimized for Aspergillus nidulans can be used for CRISPR based gene editing in filamentous fungi. We have developed a vector-based setup for Cpf1-mediated CRISPR experiments and showed that it works efficiently at different loci in A. nidulans and in A. niger. Specifically, we used our setup to demonstrate that Cpf1 is able to catalyze oligonucleotide-mediated genomic site-directed mutagenesis and marker-free gene targeting.

Conclusions
In this paper we introduce Cpf1 as a new tool in the fungal CRISPR toolbox. Our experiments demonstrate that Cpf1 can be efficiently used in Aspergilli for gene editing thereby expanding the range of genomic DNA sequences that can be targeted by CRISPR technologies.
Original languageEnglish
JournalFungal Biology and Biotechnology
Volume6
Issue number6
Number of pages10
ISSN2054-3085
DOIs
Publication statusPublished - 2019
CitationsWeb of Science® Times Cited: No match on DOI

    Research areas

  • CRISPR, Cpf1, Gene targeting, Gene editing, Filamentous fungi, Aspergilius

Download statistics

No data available

ID: 176654547