The study was done to clone and express gene of MdDpt I ( Musca domestica Diptericin I ) being screened from SSH library by salmonella, in order to lay the foundation for its bioactivity evaluation. Using cDNA end rapid amlification technology( RACE) , MdDpt I gene’ s full-length cDNA was amplified, then recombonant expression plasmid was reconstructed using pET-28a, transformed into E.coli BL21 (DE3)and then induced by IPTG, the recombinant protein was verified by SDS-PAGE. With the aid of affinity to obtain purified fusion protein, the fusion protein was verified. The full-length cDNA of MdDpt I gene was 419 bp, in size, the open reading frame was 300 bp, encoding a 99-aminoacid protein, the homology of this gene’ s cDNA sequence with the MdDpt I from GenBank was up to 95%. The mature peptide recombonant expression plasmid pET-28a-MdDpt I of MdDpt I gene was constructed, and the protein of MdDpt I gene was expressed successfully with the size of 12 ku, The purified fusion protein showed certain antibacterial activity. The result showed that the recombinant plasmid pET-28a-MdDpt I was constructed correctly, which paved the way for further studies on the MdDpt I protein expression and its biological activities.

Musca domestica L. (Diptera: Muscidae), the housefly, exhibits unique immune defenses and can produce antibacterial substances upon stimulation with bacteria. On the basis of the cDNA library constructed using the suppression subtractive hybridization method, a 1188-bp antibacterial substance gene, which we named AS566, was amplified by rapid amplification of cDNA ends from M. domestica larva stimulated with Salmonella pullorum (Enterobacteriaceae: Salmonella). In this study, the full-length AS566 gene was cloned and inserted into a His-tagged Escherichia coli (Enterobacteriaceae: Escherichia) prokaryotic expression system to enable production of the recombinant protein. The recombinant AS566 protein was purified in denatured form from inclusion bodies and renatured to obtain functionally active AS566 protein. The bacteriostatic activity of the recombinant purified AS566 protein was assessed using the Oxford plate assay system and the results indicated that AS566 had antibacterial activity against six bacteria, including an E. coli clinical isolate, S. pullorum, Streptococcus bovis (Streptococcaceae: Streptococcus), Streptococcus suis, and Staphylococcus aureus (Staphylococcaceae: Staphylococcus) in vitro. The antibacterial activity of AS566 toward Gram - bacteria was two times greater than that against Gram + bacteria. The sequencing results and BLAST analysis showed that the antibacterial substance gene AS566 was not homologous to any other antibacterial substance genes in GenBank. The antibacterial mechanisms of the newly discovered AS566 protein warrant further study.

In order to get stronger antimicrobial peptides to replace antibiotic drugs,this experiment constructed subtractive cDNA libraries about Musca domestica larvae induced by Mycoplasma hyopneumonia by suppression subtractive hybridization,and got the differential gene by reverse northern blot.The results showed that,the randomly selected positive clones were all in 200-750 bp,186 differential genes fragments screened by dot blot were cloned,sequenced and homology analysis,identified 20 protein genes fragment and 21 without homologous sequence.Musca domestica larvae induced by MHP produced anti-pathogen genes and a large number of unknown genes.