Abstract
In the CRISPR-Cas systems, Cas13a is an RNA-guided RNA nuclease specifically targeting single strand RNA. We developed a Cas13a mediated CRISPR interference tool to target mRNA for gene silencing in mosquitoes. The machinery was tested in two mosquito species. A Cas13a expressing plasmid was delivered to mosquitoes by intrathoracic injection, and Cas13a transcripts were detectable at least10 days post-delivery. In Anopheles gambiae, vitellogenin gene was silenced by Vg-crRNA injection two hours post-blood meal, which was accompanied by a significant reduction in egg production. In Aedes aegypti, the α- and δ-subunits of COPI genes were silenced by a post-blood meal crRNA injection, which resulted in mortality and fragile midguts, reproducing a phenotype reported previously. Co-silencing genes simultaneously is achievable when a cocktail of target crRNAs is given. No detectable collateral cleavages of non-target transcripts were observed in the study. This study adds a programmable CRISPR tool to manipulate RNA in mosquitoes.
Footnotes
Aditi Kulkarni, aditik{at}nmsu.edu, (575) 646-5936, Wanqin Yu, ivyyu{at}nmsu.edu, (575) 646-5936, Alex S. Moon, alexmoon{at}nmsu.edu, (575) 646-5936, Ashmita Pandey, ashmita{at}nmsu.edu, (575) 646-5936, Kathryn A. Hanley, khanley{at}nmsu.edu, (575) 646-4583, Jiannong Xu, jxu{at}nmsu.edu, (575) 646-7713
Figure 6 was added to show that Cas13a does not exhibit detectable collateral cleavage activity in mosquitoes.