Disease modeling by efficient genome editing using a near PAM-less base editor in vivo

Area of Science:

• Genetics
• Molecular Biology
• Genome Engineering

Background:

• Base editors enable precise point mutations in genomes but are limited by Protospacer Adjacent Motif (PAM) requirements.
• Existing base editing technologies restrict modeling human pathological mutations in animal models due to PAM site limitations.

Purpose of the Study:

• To develop a versatile base editor with expanded PAM recognition for efficient genome editing in animal models.
• To overcome limitations of current base editors for modeling human diseases by enabling precise mutations at more genomic sites.

Main Methods:

• Utilized the CBE4max-SpRY variant, a base editor with near PAM-less recognition, for high-efficiency point mutation introduction.
• Developed a co-selection method for identifying highly edited embryos via visual screening.
• Applied the enhanced base editor to create a zebrafish model for melanoma predisposition.

Main Results:

• Demonstrated high-efficiency point mutation introduction in an animal model using the CBE4max-SpRY variant.
• Successfully performed simultaneous multi-gene editing using the near PAM-less base editor.
• Established a novel zebrafish model for melanoma predisposition through multiplexed base editing.

Conclusions:

• The CBE4max-SpRY variant significantly expands the scope of base editing applications in animal models.
• This technology facilitates the creation of more accurate animal models for human genetic diseases.
• The developed methods enable efficient and scalable genome engineering for disease modeling and research.