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Pheromone binding proteins (PBPs) are thought to play crucial roles in perception of the sex pheromones particularly in noctuid moths, but this is rarely  in vivo  evidenced due to lacking an effective technique. Here, we reported an  in vivo  functional study of PBP1 in the important lepidopteran pest  Helicoverpa armigera  ( HarmPBP1 ), by using the CRISPR/Cas9 system. Efficient and heritable mutagenesis was achieved by egg injection of mixture of Cas9-mRNA and HarmPBP1-sgRNA. The TA cloning and sequencing revealed various insertion and/or deletion (indel) mutations at the target site. Among those, one mutation resulted in a premature stop codon at the target site, which led to a highly truncated protein with only 10 amino acids. The  HarmPBP1  with this mutation would completely loss its function, and thus was used to select the homozygous mutant insects for functional analysis. The electroantennogram recording showed that the mutant male adults displayed severely impaired responses to all three sex pheromone components (Z11-16:Ald, Z9-16:Ald and Z9-14:Ald). Our study provides the first  in vivo  evidence that  HarmPBP1  plays important role in perception of female sex pheromones, and also an effective methodology for using CRISPR/Cas9 system in functional genetic study in  H. armigera  as well as other insects.

Functional characterization of PBP1 gene in Helicoverpa armigera (Lepidoptera: Noctuidae) by using the CRISPR/Cas9 system