Cloning Of Prophenoloxidase CDNA Of Helicoverpa Armigera And Effect Of Ovomermis Sinensis Parasitization On The Transcription Of This Gene Of Helicoverpa Armigera

Insect phenoloxidase (PO) participates in melanotic encapsulation, wound healing, cuticle sclerotization and pigment forming. It is converted from prophenoloxidase(PPO) by a proPO-activating proteinase (PAP).Due to the importance of PO in insect immunity, many researchers took interest in this field. But all they have done were primarily about PO activity. In the molecular aspect it was primarily about cloning of PPO gene and there was only a little report on the influence of parasitization of parasites on the PPO transcription of their hosts.The researches about the relation of PO and insect immunity we have talked about above were primarily about bee and theire hosts. But there is only a little study on the other host-parasitoid association such as Mermithidae and insects.In this paper we selected the association of Ovomermis sinensis and Helicoverpa armigera, basic research work have been done. The main experiment contents and results are as follows:1. Cloning of PPO gene: Complete PPO cDNA sequence of Helicoverpa armigera was obtained through RT-PCR and RACE, designated Ha-ppo (Genbank accession number: DQ114946) . The cDNA was 2 405 bp in length and contained an open reading frame(ORF) of 2 097 bp which encoded 698 amino acid residues. The cDNA shared the typical structural features of PPO from other insects. Such as a possible cleavage site for proteolytic activation of the enzyme, two copper binding sites and six conserved histidine residues. The deduced amino acid sequence showed a high identity to the reported sequence of PPO2 from other insects. We deduced that the PPO gene we got was Ha-ppo2.2. PPO transcription in three tissues of 5th larvae of Helicoverpa armigera: We investigated Ha-ppo2 transcription in three tissues of 5th larva of Helicoverpa armigera using RT-PCR. The results showed that PPO mRNA was expressed in integument, haemocyte and midgut of Helicoverpa armigera .3. Effect of parasitization of Ovomermis sinensis on the transcription of PPO of Helicoverpa armigera : We studied PPO transcription of Helicoverpa armigera parasited by Ovomermis sinensis using real-time fluorescent quantitative PCR. The results showed that the transcription of Helicoverpa armigera PPO showed increase after parasited by Ovomermis sinensis.In conclusion, we cloned the full-length cDNA sequence of PPO of Helicoverpa armigera and investigated the expression profile of PPO of Helicoverpa armigera parasited by Ovomermis sinensis. All these works established the molecuar biology foundation for illustrating the molecular mechanism of Ovomermis sinensis parasitization.