A new molecular technique for determining the sex of Harmonia axyridis

Abstract

We have developed a fast and efficient PCR-based technique to sex all developmental stages of the ladybird beetle Harmonia axyridis. Previously, we established a male-specific fluorescent expressing strain of H. axyridis with the piggyBac vector. FISH analyses revealed that the transgene inserted into the H. axyridis genome in the male Y-chromosome. Because only males expressed the fluorescent signal, we were able to differentiate between the sexes of this transgenic strain even in newly hatched larvae. We took advantage of this insertion and designed an inverse PCR reaction and genome walking surrounding the inserted region to identify 3,781 bp of the male Y-chromosome genomic sequence. From this male specific region, we designed a PCR-based protocol to identify sexes from all H. axyridis developmental stages.