| 孙强正,李振军,李娟,王艺婷,金东,郑霄,张晓嫒,熊衍文,叶长芸,徐建国.霍乱毒素B亚单位在乳酸乳球菌食品级表达系统中的分泌性表达[J].Chinese journal of Epidemiology,2009,30(12):1288-1291 |
| 霍乱毒素B亚单位在乳酸乳球菌食品级表达系统中的分泌性表达 |
| Secretion expression of cholera toxin Bsubunit in food-grading Lactococcus lactis expression system |
| Received:July 30, 2009 |
| DOI： |
| KeyWord: 乳酸乳球菌 霍乱毒素B亚单位 食品级载体 分泌性表达 |
| English Key Word: Lactococcus lactis Cholera toxin B subunit Food-grade vector Secretion expression |
| FundProject:中国疾病预防控制中心青年科研基金课题(2009A103) |
| Author Name | Affiliation | E-mail | | SUN Qiang-zheng | 中国疾病预防控制中心传染病预防控制所传染病预防控制国家重点实验室, 北京 102206 | xujg@public.bta.net.cn | | LI Zhen-jun | 中国疾病预防控制中心传染病预防控制所传染病预防控制国家重点实验室, 北京 102206 | | | LI Juan | 中国疾病预防控制中心传染病预防控制所传染病预防控制国家重点实验室, 北京 102206 | | | WANG Yi-ting | 中国疾病预防控制中心传染病预防控制所传染病预防控制国家重点实验室, 北京 102206 | | | JIN Dong | 中国疾病预防控制中心传染病预防控制所传染病预防控制国家重点实验室, 北京 102206 | | | ZHENG Xiao | 中国疾病预防控制中心传染病预防控制所传染病预防控制国家重点实验室, 北京 102206 | | | ZHANG Xiao-ai | 中国疾病预防控制中心传染病预防控制所传染病预防控制国家重点实验室, 北京 102206 | | | XIONG Yan-wen | 中国疾病预防控制中心传染病预防控制所传染病预防控制国家重点实验室, 北京 102206 | | | YE Chang-yun | 中国疾病预防控制中心传染病预防控制所传染病预防控制国家重点实验室, 北京 102206 | | | XU Jian-guo |
State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China
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| Abstract: |
| 目的 在乳酸乳球菌(Lactococcus lactis)食品级分泌性表达系统L.lactis MBP71/pSQZ和L.lactis MBP71/pSQ中克隆表达霍乱毒素B亚单位(CTB),实现CTB的分泌性表达.方法 采用PCR扩增霍乱弧菌569B菌株染色体中CTB蛋白编码基因片段,克隆到食品级分泌表达载体pSQZ和pSQ中,转化入宿主菌L.lactis MBP71,构建CTB的分泌性表达系统L.lactis MBP71/pSQZ-ctB和L.lactis MBP71/pSQ-ctB;采用Western-blot检测重组菌株中CTB的表达及表达量.结果 PCR扩增得到CTB编码基因片段ctB,酶切、连接到载体pSQZ和pSQ并转化到宿主菌L.lactis MBP71中,构建了分泌性表达系统L.lactis MBP71/pSQZ-ctB和L.lactis MBP71/pSQ-ctB.Western-blot检测发现,L.lactis MBP71/pSQ-ctB和L.lactis MBP71/pSQZ-ctB系统均能分泌性表达CTB,L.lactis MBP71/pSQ-ctB上清的表达量约为2 μg/ml;L.lactis MBP71/pSQ-ctB的表达效率远远高于L.lactis MBP71/pSQZ-ctB.结论 成功实现了CTB在食品级表达系统中的分泌性表达. |
| English Abstract: |
| Objective To clone and secretion express cholera toxin B subunit(CTB)in food-grading Lactococcus lactis expression systems.Methods ctB fragment that encoding CTB was amplified by polymerase chain reaction(PCR)using the genomic DNA of Vibrio cholera strain 569B as template and was inserted into two secretion expression vector pSQZ and pSQ to construct food-grading expression system L.lactis MBP71/pSQZ-ctB and L.lactis MBP71/pSQ-ctB.The expressed CTB was detected by Western-blot assay.Results The ctB fragment was successfully amplified from Vibrio cholera strain 569B and inserted into two secretion expression vectors pSQZ and pSQ to construct food-grading expression system L.lactis MBP71/pSQZ-ctB and L.lactis MBP71/pSQ-ctB.Western-blot assay demonstrated that CTB was secretion and expressed from L.lactis MBP71 harboring vectors pSQZ-ctB and pSQ-ctB,and the quantity of CTB secreted by L.lactis MBP71/pSQ-ctB was about 2 μg/ml,higher than that of L.lactis MBP71/pSQZ-ctB.Conclusion CTB was successfully secreted and expressed by food-grading L.lactis expression systems. |
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