CRISPR Gene Drive Efficiency and Resistance Rate Is Highly Heritable with No Common Genetic Loci of Large Effect

Abstract

Gene drives could allow for control of vector-borne diseases by directly suppressing vector populations or spreading genetic payloads designed to reduce pathogen transmission. Clustered regularly interspaced short palindromic repeat (CRISPR) homing gene drives work by cleaving wild-type alleles, which are then converted to drive alleles by homology-directed repair, increasing the frequency of the drive in a population over time. However, resistance alleles can form when end-joining repair takes place in lieu of homology-directed repair. Such alleles cannot be converted to drive alleles, which would eventually halt the spread of a drive through a population. To investigate the effects of natural genetic variation on resistance formation, we developed a CRISPR homing gene drive in Drosophila melanogaster and crossed it into the genetically diverse Drosophila Genetic Reference Panel (DGRP) lines, measuring several performance parameters. Most strikingly, resistance allele formation postfertilization in the early embryo ranged from 7 to 79% among lines and averaged 42 6 18%. We performed a genome-wide association study using our results in the DGRP lines, and found that the resistance and conversion rates were not explained by common alleles of large effect, but instead there were several genetic polymorphisms showing weak association. RNA interference knockdown of several genes containing these polymorphisms confirmed their effect, but the small effect sizes imply that their manipulation would likely yield only modest improvements to the efficacy of gene drives.

Performance analysis of novel toxin-antidote CRISPR gene drive systems

“…Complete suppression occurs when the suppression factor is greater than the maximum population growth rate experienced by individuals at lower densities with less competition, before Allee effects dominate 32 . For the germline cut rates that have been observed experimentally in mosquito and Drosophila systems [12][13][14][15][16][17][18][19][20][21] , this would likely be sufficient to cause complete suppression. High suppression factors are also possible even if the target gene shows only partial haploinsufficiency ( Figure S4).…”

“…Homing gene drive constructs based on CRISPR-Cas9 have been constructed in yeast [8][9][10][11] , flies [12][13][14][15][16][17][18] , mosquitoes [19][20][21] , and mice 22 . These constructs work by cleaving a wild-type allele, resulting in copying of the drive allele during homology-directed repair.…”

Computational and experimental performance of CRISPR homing gene drive strategies with multiplexed gRNAs

“…The drive is then copied into the wild-type site via homology-directed repair, increasing the frequency of the drive allele in the population. Thus far, CRISPR homing gene drives have been demonstrated in yeast [5][6][7][8] , flies [9][10][11][12][13][14][15][16] , mosquitoes [17][18][19] , and mice 20 .…”

A toxin-antidote CRISPR gene drive system for regional population modification

“…CRISPR-based homing drives promise a flexible gene drive mechanism for both population modification and suppression, and such systems have now been demonstrated in a variety of organisms, including yeast [8][9][10][11] , flies [12][13][14][15][16][17][18] , mosquitoes [19][20][21] , and mice 22 . These constructs work by cleaving a wild-type allele at a predetermined target site.…”