中国兽医科学2012，Vol.42Issue(4)：368-372,5.
副鸡嗜血杆菌基因文库的构建与筛选
Construction and screening of genomic library of Haemophilus paragallinarum
摘要
Abstract
In order to select in vivo expression genes of Haemophilus paragallinarum,the total DNA of the bacterium was extracted and the genomic library was constructed with pET system.The positive clones in the library were identified by PCR and SalⅠ-NdeⅠ-digestion.The expressed library was screened by colony hybridization using rehabilitation serum,which was absorbed with H.paragallinarum,as the probe.In result,the recombinant plasmids contained from 0.5 to 2 kb of target fragments,and 99% of H.paragallinarum genes were involved in the library.The 17 positive clones obtained by colony hybridization were confirmed by PCR and SalⅠ-NdeⅠ digestion followed by direct-sequencing.Sequences were then blasted in the NCBI GenBank.The sequence analysis revealed 5 ORFs encoded glutamyl tRNA reductase,transcription termination factor,capsule biosynthesis region 2 gene cluster,and two hypothetical proteins,respectively.The results showed that the study identified some in vivo induced genes of H.paragallinarum by IVIAT approach and did some initial work about the gene function.We marched one step on the road of exploring the key genes which help H.paragallinarum survive in the body and result in infectious coryza.And the study also provided some new molecular targets data of prophylaxis and therapy of infectious coryza.关键词
副鸡嗜血杆菌/基因文库构建/基因文库筛选/菌落原位杂交Key words
Haemophilus paragallinarum/genomic library construction/genomic library screening/colony hybridization分类
农业科技引用本文复制引用
王雪敏,梁玉荣,吕学泽,张培君,龚玉梅,王宏俊..副鸡嗜血杆菌基因文库的构建与筛选[J].中国兽医科学,2012,42(4):368-372,5.基金项目
国家“十二五”高技术研究发展计划（863）项目 （）
北京市自然科学基金资助项目 （）
北京市优秀人才培养资助项目 （）
