Upper bound on the mutational burden imposed by a CRISPR-Cas9 gene-drive element

Area of Science:

• Genetics
• Molecular Biology
• Evolutionary Biology

Background:

• CRISPR-Cas9 gene drives (CCGDs) offer potential for population genetic control but raise safety concerns.
• Off-target DNA breaks by Cas9 could increase mutation rates and cause loss-of-heterozygosity (LOH).
• The magnitude of these unintended effects remains largely unknown.

Purpose of the Study:

• To quantify the impact of CCGDs and Cas9 on mutation and LOH rates.
• To assess the long-term evolutionary safety of gene drive systems.

Main Methods:

• Mutation accumulation experiment in yeast (Saccharomyces cerevisiae).
• Genome-wide analysis of de novo mutations and LOH events.
• Power calculations to determine effect size detection limits.

Main Results:

• No significant increase in genome-wide mutation or LOH rates was detected with CCGD or Cas9 presence.
• Power analysis indicated effects, if any, were less than 30%, below natural yeast variation.
• Potential alterations in LOH event characteristics were observed but lacked strong statistical support.

Conclusions:

• CCGDs impose a minimal additional mutational burden in yeast.
• Gene drive systems may present acceptable evolutionary risks.
• Further evaluation in diverse systems is recommended to confirm safety.