1.Study On Expression And Function Of WAP Antibacterial Peptide 2.Identification And Expression Analysis Of Key Genes In Housefly ProPO System

Invertebrates do not have acquired immunity, only have innate immunity, wich including cellular immunity and humoral immunity. antibacterial peptide and proPO activation system are two important of components humoral immunity, and play an important role in the insect’s immune system. In this paper, we mainly studied the WAP antimicrobial peptides and propo activation system, Especially the function of antimicrobial peptides in eukaryotic organisms and it’s expression in vitro. I still studied the location and classification of key genes in the propo activation system, and made a preliminary exploration of their expression mode.Useing the Drosophila,which belongs to eukaryotes and have clear genetic background,we import exogenous WAP gene into Drosophila’s body, and built the UAS-wap turn of a Drosophila strains. Using the genetic variation method i finished the gene mapping of wap’s chromosomal location of UAS-wap transgenic Drosophila.I found that the wap were inserted in sex chromosomes,chromosome second,chromosome third of transgenic Drosophila,and let them hybridize With actin-Gal4 Drosophila to realize the overexpression of WAP gene in their hybrid progeny.Found by qPCR, WAP gene expression at mRNA level was significantly higher than that of control, some up to 100 times.To inquiry and verification the antibacterial function of WAP antibacterial peptide in transgenic Drosophila,it accessed to the specific antimicrobial effect. The results laid a good foundation for the development of newantibacterial drugs.With the attention degree of antimicrobial peptides increasingly high,People are already successfully obtained the recombinant protein of WAP using the prokaryotic expression system in vitro,and measured it has obvious antibacterial activity,But the expression of WAP antibacterial peptide in eukaryotic system in vitro have not been successful.This paper did a research about WAP antibacterial peptide’s expression in eukaryotic system,and successfully constructed the recombinant expression vector of wap/pPIC9K,added we obtained the eukaryotic expression strain which containing wap/pPIC9K recombinant plasme.In addition to the technical problem that the WAP antimicrobial peptides expressioned by prokaryotic system are exist in the form of inclusion body,which is not conducive to the WAP to play biological activity and to be used.We want to find a method which can obtain high yield of soluble WAP,so we expressed WAP in prokaryotic expression system.we successfully built the wap/pET32a/Rosetta recombinant expression strain,obtained the soluble recombinant protein-WAP peptide by Induced expression in vitro.That is a major technological breakthrough,and also laid the technical foundation for the early realization of industrialized production of WAP antibacterial peptide.At the same time,another major factor-proPO activation system-in humoral immunity,has gradually attracted the attention of science field,now At home and abroad are still not clear about all components of proPO activation system,we foucs on studied several key genes in the proPO activation system:Prophenoloxidase(proPO),Activation of the enzyme(PAP).Inhibitory factor(Serpin),Factor receptor(PGRP).we identified and classified these key genes,and analyzed their expression patterns.In this paper,through the transcriptome sequencing,sequence analysis and gene cloning,we screened gene Unigeens of Serpins class and PAP class,and after sequence assembly,Verification,analysis and prediction of structure and function;we initially divided Serpins class into 11 categories,with the PAP class was divided into 3 categories.Added we analyzed the Serpin’s expression pattern at different developmental stages in housefly larvae,and a preliminary study on the PAP’s vivo function in housefly larvae.This study can provide for scientific information about the mechanism of activation and immune function of housefly proPO system,and it has important theoretical and practical significance To reveal the unique and effective immune mechanism of housefly;find the key immune function molecular,and development a new method of effectivly controled the pest and serious animal infectious diseases.