CRISPR/Cas Genome Editing in Plant Fungi

Completing sequencing an increasing number of fungal genomes provides the conditions for revealing the function of fungal genes at the analytical level. However, the study of fungal genetics is seriously hampered by the relatively large fungal genomes, complex cell structure, lack of sexual stages in most fungi, and difficulties in gene manipulation. Lifeasible can establish fungal genetic transformation systems that provide for targeted genetic modification of fungal strains through insertion, deletion, and knock-down strategies.

Figure 1. The history of the development and application of CRISPR/Cas9 technology in filamentous fungi. (Song R, et al., 2019)

The CRISPR/Cas system consists of a manipulator encoding a Cas protein and a CRISPR repeat spacer sequence. A typical CRISPR repeat spacer sequence consists of a series of short, highly conserved forward repeats and spacer sequences of similar lengths in a certain order. Cas genes are a conserved family of genes that encode nucleases, DNA-decapping enzymes, polymerases, and other proteins related to the cleavage and modification of nucleic acids.

Genetic manipulation in fungi is often time-consuming and cumbersome, especially in diploid strains that lack a sexual cycle. Two core components of the CRISPR/Cas gene editing system include the Cas9 protein, which is used to generate DNA double-strand breaks, and the sgRNA, which is capable of directing the Cas9 nucleic acid endonuclease to any locus in a chromosome with a suitable prototypical spacer sequence adjacent to the motif.

The mechanism of action of CRISPR/Cas-mediated fungal gene editing consists of three main steps, which are the acquisition of new spacer sequences, tracrRNA synthesis and processing, and CRISPR/Cas9-mediated silent immune interference. When the constructed exogenous CRISPR/Cas system is introduced into the recipient cell, the sgRNA recognizes the target site, and Cas9 is responsible for the cleavage of the fungal DNA double strand; we can help our customers to introduce the target DNA fragment into the cell and realize the targeted editing of the target gene.

• Filamentous fungal gene editing. Filamentous fungi are widely distributed in nature and are closely related to human production and life. Many filamentous fungal genomes have been sequenced, and the study of filamentous fungi has entered the post-genomic era. We can provide gene editing services for filamentous fungi, including gene insertion, deletion and base conversion, and transcriptional activation, through the CRISPR-Cas system. CRISPR/Cas9-mediated gene editing in filamentous fungi mainly includes the following strains, Aspergillus, Trichoderma reesei, Ustilagomaydis, and Pyriculariaoryzae.
• Gene editing of plant pathogenic fungi. We can design sgRNAs based on target gene sequences by CRISPR/Cas9-mediated genome editing technology to achieve targeted editing of pathogenic fungal genomes and obtain plant pathogenic strains with target gene editing mutations through resistance and molecular biology screening.

Lifeasible can use CRISPR/Cas9 gene editing systems to help customers advance their research related to fungal functional gene mining, targeted improvement of varieties, and targeted mutation of development-related genes. As your trusted partner, we can meet all your fungal phylogenetic analysis needs and provide you with efficient and high-quality services. If you want to know the details, please contact us.