藏鸡源乳酸菌的筛选鉴定及IBV N基因的表达研究
Isolation and identification of lacticacid bacteria from Tibetan chicken and expression of foreign protein in it
作者：刘思彤(四川大学生命科学学院，动物疫病防控与食品安全四川省重点实验室；生物资源与生态环境教育部重点实验室)；王红宁(四川大学生命科学学院，动物疫病防控与食品安全四川省重点实验室；生物资源与生态环境教育部重点实验室)；马炳存(四川大学生命科学学院，动物疫病防控与食品安全四川省重点实验室；生物资源与生态环境教育部重点实验室)；丁梦蝶(四川大学生命科学学院，动物疫病防控与食品安全四川省重点实验室；生物资源与生态环境教育部重点实验室)；曹海鹏(四川大学生命科学学院，动物疫病防控与食品安全四川省重点实验室；生物资源与生态环境教育部重点实验室)；杨鑫(四川大学生命科学学院，动物疫病防控与食品安全四川省重点实验室；生物资源与生态环境教育部重点实验室)
Author：LIU Si-Tong(Animal Disease Prevention and Food Safety Key Lab of Sichuan Province, Key Laboratory of Bio resources and Eco environment of Ministry of Education, College of Life Sciences, Sichuan University)；WANG Hong-Ning(Animal Disease Prevention and Food Safety Key Lab of Sichuan Province, Key Laboratory of Bio resources and Eco environment of Ministry of Education, College of Life Sciences, Sichuan University)；MA Bing-Cun(Animal Disease Prevention and Food Safety Key Lab of Sichuan Province, Key Laboratory of Bio resources and Eco environment of Ministry of Education, College of Life Sciences, Sichuan University)；DING Meng-Die(Animal Disease Prevention and Food Safety Key Lab of Sichuan Province, Key Laboratory of Bio resources and Eco environment of Ministry of Education, College of Life Sciences, Sichuan University)；CAO Hai-Peng(Animal Disease Prevention and Food Safety Key Lab of Sichuan Province, Key Laboratory of Bio resources and Eco environment of Ministry of Education, College of Life Sciences, Sichuan University)；YANG Xin(Animal Disease Prevention and Food Safety Key Lab of Sichuan Province, Key Laboratory of Bio resources and Eco environment of Ministry of Education, College of Life Sciences, Sichuan University)
收稿日期：2014-04-05 年卷（期）页码：2015,52(3):677-681
期刊名称：四川大学学报: 自然科学版
Journal Name：Journal of Sichuan University (Natural Science Edition)
关键字：藏鸡; 乳酸菌; IBV N基因; 表达
Key words：Tibetan chicken; Lactobacillus; IBV N gene; expression
基金项目：国家“863”计划（2011AA10A209）； 现代农业产业技术体系建设专项资金项目（CARS-41-K09）； 四川省应用基础研究计划项目(2013JY0027)； 四川省科技基础条件平台项目（14010136）； 四川省科技支撑计划项目（2014NZ0020）
中文摘要
从藏鸡(海拔3000米藏区、自然放养、未使用过抗生素)肠道内筛选乳杆菌, 经16s rDNA和生理生化试验鉴定为植物乳杆菌.通过耐酸耐胆汁试验和肠道黏附试验测试, 表明其具有良好的耐酸耐胆汁能力和肠道黏附能力.为使外源质粒顺利转化入乳杆菌, 进行电场强度的优化, 再按照优化后的电场强度, 将禽冠状病毒pSIP409 N重组质粒电转化入该株乳杆菌, 进行双酶切和PCR鉴定.将经鉴定成功转入的重组乳杆菌, 用诱导肽SppIP诱导表达, 再进行SDS PAGE检测外源蛋白的表达.结果表明N蛋白能够顺利表达. 上述试验证明该株植物乳杆菌具有作为乳酸菌活载体疫苗的潜力.
英文摘要
One lactobacillus strain was isolated from free ranging Tibetan chicken in Ganzi autonomous county, Sichuan Province. The strain was identified as Lactobacillus plantarum by 16s rDNA sequence and physiological and biochemical identification. The tests for growth with bile salts, tolerance to acidic pH, and adhesion efficiency to the epithelial cells of chickens were also carried out. These data suggested the strain has a good ability to survive in gastrointestinal tract and a high adhesion efficiency to the epithelial cells. We then estimated the isolated lactobacillus as a host to express foreign genes. Recombinant plasmid pSIP409 N inserted nucleoprotein (N) of IB virus was electroporated into the strain, and the N protein was successfully expressed while it was induced by SppIP. These results indicated that the foreign recombinant plasmid can be transformed into L.plantarum and expressed efficiently. Conclusively, the present study demonstrated that L.plantarum is a potential live vector strain for poultry vaccine.
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