A platform for precise quantification of gene editing products based on microfluidic chip-based digital PCR

Area of Science:

• Biotechnology
• Molecular Biology
• Genomics

Background:

• CRISPR/Cas9 and derivatives enable precise gene editing.
• Low editing efficiency poses challenges for detecting and quantifying gene editing products.
• Existing methods lack precise quantification capabilities.

Purpose of the Study:

• To develop a novel platform for precise quantification of gene editing products.
• To assess gene editing efficiency with high sensitivity and accuracy.
• To overcome limitations in detecting low-level gene editing events.

Main Methods:

• Development of a microfluidic chip-based digital polymerase chain reaction (dPCR) assay.
• Application of the platform for quantifying gene editing products in various organisms.
• Utilizing dPCR for absolute quantification of nucleic acid samples.

Main Results:

• The platform accurately identified different types of edited samples, including complex genomes like tetraploid rapeseed and soybean.
• Achieved high sensitivity as low as 8.14 copies/μL and a detection limit of 0.1%.
• Demonstrated superior performance with high sensitivity, low detection limit, and wide applicability.

Conclusions:

• Microfluidic chip-based dPCR is a powerful tool for precise quantification of gene editing products.
• The developed platform enables accurate assessment and quantification of gene editing efficiency.
• This technology addresses the need for precise analysis of low-level gene editing events.