Given the super-Mendelian inheritance achieved by CRISPR-Cas9 homing gene drives, this technology could help modify or eradicate pests and disease-carrying insects. For instance, in the case of disease vectors such as mosquitoes, CRISPR gene drives may be used to disseminate—through populations—transgenes that prevent disease transmission. This technology relies on the activity of Cas9 endonuclease in the germline, where it induces—at the target site—chromosomal breaks. For drive conversion to occur, these breaks must be resolved by the homology-directed repair pathway (HDR). As HDR predominates specifically during early meiosis, Cas9-induced breaks should occur during early meiosis. When the breaks are produced and resolved before or after the window optimal for HDR, nonhomologous end joining (NHEJ) prevails and resistance alleles, which impede drive conversion, are produced. Therefore, the ideal gene drive cassette would restrict Cas9 expression not only spatially, to the germline, but also temporally, to early meiosis.

 

However, even using promoters that are only active during pre-meiosis, mRNA and proteins can persist in the egg until advanced embryogenesis. Therefore, a drive-carrying female typically deposits nuclease protein or mRNA in the egg. Then, premature cleavage by this maternally deposited nuclease in the resulting embryo has been the major source of end-joining mutations and resistance alleles in current gene drive designs.

​

Future research focused on silencing Cas9 during or after meiosis to restrict nuclease activity to the early meiosis will possibly benefit the development of this technology. Studying gene expression dynamics of pre-, early, and late meiosis may help identify cis-regulatory elements that could be used for RNAi targeting Cas9 mRNA. Gene drives carrying a Cas9-RNA interference construct that knocks down Cas9 transcript at the beginning of meiosis may exhibit reduced levels of Cas9 protein during late oogenesis and embryogenesis, bringing down resistance allele formation.

​