Optimization of the base editor BE4max in chicken somatic cells

Area of Science:

• Animal reproductive and breeding biotechnology
• Gene editing technologies
• Poultry genetics

Background:

• Gene editing allows trait alteration and genetic resource creation in livestock.
• Base editing (BEs) enables single-base mutations without DNA breaks, showing promise for livestock genetic modification.
• Existing base editors require optimization for specific applications, such as in chicken somatic cells.

Purpose of the Study:

• To evaluate and optimize the efficacy of the BE4max base editor in chicken somatic cells (DF-1).
• To improve base editing efficiency for potential applications in poultry breeding.

Main Methods:

• Optimized the cytosine deaminase (APOBEC) component of BE4max for chicken.
• Tested the optimized chicken base editor (chBE4max) in DF-1 cells.
• Inhibited methyl binding domain protein 4 (MBD4) expression using siRNA to assess its impact on editing efficiency.

Main Results:

• The optimized chBE4max editor showed a 10.4% ± 4.6 improvement in base editing efficiency compared to the original BE4max.
• Inhibiting MBD4 gene expression via siRNA further enhanced editing efficiency by 4.43% ± 1.4 in chicken DF-1 cells.
• Demonstrated successful optimization of base editing tools for chicken genetic applications.

Conclusions:

• The optimized chBE4max editor significantly improves base editing efficiency in chicken somatic cells.
• Targeted inhibition of MBD4 can further enhance base editing outcomes.
• This optimized base editing technology holds potential for advancing poultry breeding studies and genetic resource development.