Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2025:2892:49-67.
doi: 10.1007/978-1-0716-4330-3_4.

Gene Editing and Protein Tagging in the Oomycete Phytophthora infestans Using CRISPR-Cas12a

Carl S Mendoza  1 , Audrey M V Ah-Fong  1 , Howard S Judelson  2
Affiliations

Gene Editing and Protein Tagging in the Oomycete Phytophthora infestans Using CRISPR-Cas12a

Carl S Mendoza et al. Methods Mol Biol. 2025.

Abstract

Molecular genetic tools such as CRISPR-Cas gene editing systems are invaluable for understanding gene and protein function and revealing the details of a pathogen's life and disease cycles. Here we present protocols for genome editing in Phytophthora infestans, an oomycete with global importance as a pathogen of potato and tomato. Using a vector system that expresses variants of Cas12a from Lachnospiraceae bacterium and its guide RNA from a unified transcript, we first present a method for editing genes through the non-homologous end-joining (NHEJ) pathway. We then describe an application of homology-directed repair (HDR), in which Cas12a is used to fuse a protein-coding gene with a fluorescent or epitope tag. Both methods should be adaptable to many oomycetes other than P. infestans.

Keywords: CRISPR; Fluorescent reporter; Gene editing; Oomycete; Transformation.

PubMed Disclaimer

References

    1. Anzalone AV, Koblan LW, Liu DR (2020) Genome editing with CRISPR-Cas nucleases, base editors, transposases and prime editors. Nat Biotechnol 38:824–844 - DOI - PubMed
    1. Fang Y, Tyler BM (2016) Efficient disruption and replacement of an effector gene in the oomycete Phytophthora sojae using CRISPR/Cas9. Mol Plant Pathol 17:127–139 - DOI - PubMed
    1. Ah-Fong A, Boyd AM, Matson MEH, Judelson HS (2021) A Cas12a-based gene editing system for Phytophthora infestans reveals monoallelic expression of an elicitor. Mol Plant Pathol 22:737–752 - DOI - PubMed - PMC
    1. Gumtow R, Wu DL, Uchida J, Tian MY (2018) A Phytophthora palmivora extracellular cystatin-like protease inhibitor targets papain to contribute to virulence on papaya. Mol Plant-Microbe Interact 31:363–373 - DOI - PubMed
    1. Chen XR, Zhang Y, Li HY, Zhang ZH, Sheng GL, Li YP et al (2019) The RXLR effector PcAvh1 is required for full virulence of Phytophthora capsici. Mol Plant-Microbe Interact 32:986–1000 - DOI - PubMed

MeSH terms

LinkOut - more resources