A CRISPR homing gene drive targeting a haplolethal gene removes resistance alleles and successfully spreads through a cage population

J. Champer, E. Yang, E. Lee, J. Liu, A. G. Clark and P. W. Messer,  Proceedings of the National Academy of Sciences,  202004373. 2020.

CRISPR gene drives have sparked considerable interest among scientists and the public alike due to their potential for genetic alteration of entire species. However, despite the initial hype, experiments soon revealed that such drives suffer from a critical flaw—the rapid evolution of resistance alleles. These alleles are frequently formed when CRISPR-induced cleavage is repaired by end joining and will hinder the spread of the drive. Here, we present a CRISPR homing drive that was able to successfully spread to all individuals in a laboratory cage study in Drosophila melanogaster without any apparent evolution of resistance. The drive achieves this by targeting a haplolethal gene, resulting in individuals with resistance alleles being nonviable due to disruption of the target gene.Engineered gene drives are being explored as a new strategy in the fight against vector-borne diseases due to their potential for rapidly spreading genetic modifications through a population. However, CRISPR-based homing gene drives proposed for this purpose have faced a major obstacle in the formation of resistance alleles that prevent Cas9 cleavage. Here, we present a homing drive in Drosophila melanogaster that reduces the prevalence of resistance alleles below detectable levels by targeting a haplolethal gene with two guide RNAs (gRNAs) while also providing a rescue allele. Resistance alleles that form by end-joining repair typically disrupt the haplolethal target gene and are thus removed from the population because individuals that carry them are nonviable. We demonstrate that our drive is highly efficient, with 91% of the progeny of drive heterozygotes inheriting the drive allele and with no functional resistance alleles observed in the remainder. In a large cage experiment, the drive allele successfully spread to all individuals within a few generations. These results show that a haplolethal homing drive can provide an effective tool for targeted genetic modification of entire populations.All study data are included in the article and supporting information.

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