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Title: Double-tap gene drive uses iterative genome targeting to help overcome resistance alleles
Abstract

Homing CRISPR gene drives could aid in curbing the spread of vector-borne diseases and controlling crop pest and invasive species populations due to an inheritance rate that surpasses Mendelian laws. However, this technology suffers from resistance alleles formed when the drive-induced DNA break is repaired by error-prone pathways, which creates mutations that disrupt the gRNA recognition sequence and prevent further gene-drive propagation. Here, we attempt to counteract this by encoding additional gRNAs that target the most commonly generated resistance alleles into the gene drive, allowing a second opportunity at gene-drive conversion. Our presented “double-tap” strategy improved drive efficiency by recycling resistance alleles. The double-tap drive also efficiently spreads in caged populations, outperforming the control drive. Overall, this double-tap strategy can be readily implemented in any CRISPR-based gene drive to improve performance, and similar approaches could benefit other systems suffering from low HDR frequencies, such as mammalian cells or mouse germline transformations.

 
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Award ID(s):
2048207
NSF-PAR ID:
10366756
Author(s) / Creator(s):
; ; ; ; ;
Publisher / Repository:
Nature Publishing Group
Date Published:
Journal Name:
Nature Communications
Volume:
13
Issue:
1
ISSN:
2041-1723
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
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