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Updated: Jun 9, 2025
CRISPR-mediated Genome Editing of the Human Fungal Pathogen Candida albicans
Published on: November 14, 2018
CRISPR-GRIT: Guide RNAs with Integrated Repair Templates Enable Precise Multiplexed Genome Editing in the Diploid
Christopher J Cotter1, Cong T Trinh1
1Department of Chemical and Biomolecular Engineering, The University of Tennessee, Knoxville, Tennessee, USA.
Abstract:
Candida albicans, an opportunistic fungal pathogen, causes severe infections in immunocompromised individuals. Limited classes and overuse of current antifungals have led to the rapid emergence of antifungal resistance. Thus, there is an urgent need to understand fungal pathogen genetics to develop new antifungal strategies. Genetic manipulation of C. albicans is encumbered by its diploid chromosomes requiring editing both alleles to elucidate gene function. Although the recent development of CRISPR-Cas systems has facilitated genome editing in C. albicans, large-scale and multiplexed functional genomic studies are still hindered by the necessity of cotransforming repair templates for homozygous knockouts. Here, we present CRISPR-GRIT (Guide RNAs with Integrated Repair Templates), a repair template-integrated guide RNA design for expedited gene knockouts and multiplexed gene editing in C. albicans. We envision that this method can be used for high-throughput library screens and identification of synthetic lethal pairs in both C. albicans and other diploid organisms with strong homologous recombination machinery.
Insights
A new CRISPR-GRIT tool simplifies gene editing in Candida albicans, accelerating the development of novel antifungal strategies against resistant fungal infections.
Area of Science:
- Mycology
- Molecular Biology
- Genetics
Background:
- * *Candida albicans* is an opportunistic fungal pathogen causing severe infections, particularly in immunocompromised individuals.
- * Antifungal resistance is rapidly emerging due to limited drug classes and overuse of current treatments.
- * Understanding fungal genetics is crucial for developing new antifungal therapies, but genetic manipulation in diploid *C. albicans* is challenging.
Purpose of the Study:
- * To develop an expedited method for gene knockouts and multiplexed gene editing in *Candida albicans*.
- * To overcome limitations in large-scale functional genomic studies caused by the need for cotransforming repair templates for homozygous knockouts.
Main Methods:
- * Introduction of CRISPR-GRIT (Guide RNAs with Integrated Repair Templates).
- * A novel repair template-integrated guide RNA design for efficient gene editing.
- * Application in *Candida albicans*, a diploid organism with strong homologous recombination.
Main Results:
- * CRISPR-GRIT enables expedited gene knockouts.
- * The method facilitates multiplexed gene editing in *C. albicans*.
- * Demonstrates a streamlined approach for genetic manipulation in diploid organisms.
Conclusions:
- * CRISPR-GRIT significantly enhances the efficiency of gene editing in *Candida albicans*.
- * This method is suitable for high-throughput library screens and identifying synthetic lethal pairs.
- * CRISPR-GRIT offers a valuable tool for advancing antifungal research and developing new therapeutic strategies.
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