JOURNAL ARTICLE
RESEARCH SUPPORT, NON-U.S. GOV'T
Add like
Add dislike
Add to saved papers

Design, execution, and analysis of CRISPR-Cas9-based deletions and genetic interaction networks in the fungal pathogen Candida albicans.

Nature Protocols 2019 March
The study of fungal pathogens is of immediate importance, yet progress is hindered by the technical challenges of genetic manipulation. For Candida species, their inability to maintain plasmids, unusual codon usage, and inefficient homologous recombination are among the obstacles limiting efficient genetic manipulation. New advances in genomic biotechnologies-particularly CRISPR-based tools-have revolutionized genome editing for many fungal species. Here, we present a protocol for CRISPR-Cas9-based manipulation in Candida albicans using a modified gene-drive-based strategy that takes ~1 month to complete. We detail the generation of Candida-optimized Cas9-based plasmids for gene deletion, an efficient transformation protocol using C. albicans haploids, and an optimized mating strategy to generate homozygous single- and double-gene diploid mutants. We further describe protocols for quantifying cell growth and analysis pipelines to calculate fitness and genetic interaction scores for genetic mutants. This protocol overcomes previous limitations associated with genetic manipulation in C. albicans and advances researchers' ability to perform genetic analysis in this pathogen; the protocol also has broad applicability to other mating-competent microorganisms.

Full text links

We have located links that may give you full text access.
Can't access the paper?
Try logging in through your university/institutional subscription. For a smoother one-click institutional access experience, please use our mobile app.

For the best experience, use the Read mobile app

Mobile app image

Get seemless 1-tap access through your institution/university

For the best experience, use the Read mobile app

All material on this website is protected by copyright, Copyright © 1994-2024 by WebMD LLC.
This website also contains material copyrighted by 3rd parties.

By using this service, you agree to our terms of use and privacy policy.

Your Privacy Choices Toggle icon

You can now claim free CME credits for this literature searchClaim now

Get seemless 1-tap access through your institution/university

For the best experience, use the Read mobile app