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suicide vector Construction of Haemophilus parasuis hhd B Gene Marker-free Deleted

Animal Husbandry and Feed Science 2015年第6期7卷 354-357页

作者： Song Shuai Li Miao Li Yan Jiang Zhiyong Cai Rujian Yang Dongxia Li Chunling Institute of Animal Health Guangdong Academy of Agricultural Sciences Guangdong Key Labratory of Animal Disease Prevention

To construct the suicide vector of hhd B gene marker-free mutant in Haemophilus parasuis( HPS),two pairs of specific primers were designed and synthesized according to the hhd B gene upstream and downstream sequences of HPS published in Gen Bank. The hhd B gene upstream and downstream sequences were amplified by PCR,which were further ligated( hhd B-up + down) through overlapping PCR method. NotⅠand SalⅠrestriction enzyme sites were introduced on both ends of the ligated sequence. After the corresponding digestion,the hhd B-up + down sequence was directionally cloned to the suicide plasmid vector p EMOC2. Results showed that the suicide vector of hhd B gene marker-free deleted( p EMOC2Δhhd B) with stable inheritance in E. coli β2155 strain was successfully obtained,thereby laying the foundation for construction of HPS-hhd B gene marker-free mutant strain.

关键词： Haemophilus parasuis(HPS) hhd B gene suicide vector

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A Non-Marker Mutagenesis Strategy to Generate Poly-hrp Gene Mutants in the Rice Pathogen Xanthomonas oryzae pv. oryzicola

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Agricultural Sciences in China 2011年第8期10卷 1139-1150页

作者： ZOU Li-fang LI Yu-rong CHEN Gong-you School of Agriculture and Biology Shanghai Jiao Tong UniversityShanghai 200240P.R.China Key Laboratory of Monitoring and Management for Plant Diseases and Insects Ministry of Agriculture/Department of Plant PathologyNanjing Agricultural UniversityNanjing 210095P.R.China

Xanthomonas oryzae *** （Xoc）,the critical pathogen causing bacterial leaf streak in rice,possesses a hrp cluster that is responsible for triggering hypersensitive response （HR） in non-host tobacco and pathogenicity in host rice,and is considered to be one of the model pathogens in the rice model ***,we developed a high-throughput mutagenesis system using a two-step integration mediated by a novel suicide vector *** was used to generate single or poly-gene mutants of hpa1,hpa2,hrcV,hrpE,hpaB,and hrpF gene for functional *** total,five single,four double,and two triple hrp gene mutants were *** double and triple hrp gene deletion mutants triggered novel phenotypes in *** data suggest that pKMS1 is a useful tool for non-marker mutagenesis of multiple genes in Xoc.

关键词： Xanthomonas oryzae pv. oryzicola suicide vector knockout mutagenesis hrp gene

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