Screening And Analysis Of In Vivo Induced Genes Of Avibacterium Paragallinarum

Avibacterium paragallinarum (Apg) is the causative agent of infectious coryza.The disease causes retarded growth of chickens and a reduction in egg production, and subsequently, serious economic losses to the poultry industry. So it is very imminent to find new drugs and vaccines to deal with the crisis.After Apg invades the host body, a series of bacterial genes will start up and express to adapt the internal environment. These in vivo induced genes may be the key ones to cause the disease and molecular targets of theimmunological and therapeutic research.Genomic DNA from Apg was extracted. The DNA was partially digested with Sau3A I and the fragments were inserted into the pET system expression vectors to construct the genomic library. Using the principle of in vivo induced antigen technology, the library was induced with IPTG and then was screened with the sera pooled from naturally infected chicikens and absorbed with in vitro grown Apg and Escherichia coli BL21(DE3). The inserts of positive clones were sequenced with primer T7promoter. And then analysis the sequences in NCBI to identify the open reading frames. To get H18capsule biosynthesis region2gene cluster as the research object construct the deletion vector, and then electrotransforme it into Apg. Transformants were selected with kanamycin and sucrose. The H18L mutagenesis strains were preliminarily identified using PCR.We constructed the genomic expression library of Apg. The library included6×103clones, and more than eighty percent were recombinated plasmids. The library met with the theoretieal requirements. After screening, sequenceing and blasting, five open reading frames were obtained in this study including one encoding glutamyl tRNA reductase, one transcription termination factor, one H18capsule biosynthesis region2gene cluster, and two hypothetical proteins. Based on this, we successfully constructed H18L gene deletion vector and electrotransformed it into Apg, This work is underway.This study identified some in vivo induced genes of Apg by IVIAT approach and then did some initial work about the gene function. We laid a foundation for exploring the key genes which help Apg survive in the body and result in infectious coryza.And the study also provided some ideas of prophylaxis and therapy of infectious coryza.