Identification of yellow gene family in Agrotis ipsilon and functional analysis of Aiyellow-y by CRISPR/Cas9

Abstract

The yellow gene family has been identified in several model insects, but yellow genes were poorly identified in non-model insects and the functions of yellow genes are largely unknown. In this study, we identified seven yellow genes in an important agricultural pest Agrotis ipsilon. Each gene encodes a protein containing a major royal jelly domain. Phylogenetic analysis defined these genes as yellow-y, -b, -b2, -c, -d, -e, and -h, respectively. The A. ipsilon yellow genes yellow-b, -b2, and -c were stably expressed in all developmental stages and tissues analyzed, whereas the other four yellow genes had unique expression patterns, suggesting distinct physiological roles of each gene. Using the CRISPR/Cas9 system, we successfully disrupted yellow-y in A. ipsilon and obtained G₀ insects with somatic mutations. Unlike the black of wild-type newly hatched larvae and of adults, the mutants were yellow, although in the pupal stage mutant coloration did not differ from wild-type coloration. This phenotype was inherited by G₁ offspring. The G₀ mutants did not show any growth deficiency compared with control insects; however, a dehydration-like phenotype was observed in newly hatched G₁ larvae from sibling crossed mutants. Our results indicate that A. ipsilon yellow-y gene plays a role in body pigmentation and also might function in waterproofing.

Keywords: Agrotis ipsilon; CRISPR/Cas9; Pigmentation; Yellow family; Yellow-y.