牙龈卟啉单胞菌菌毛融合抗原基因果实特异表达载体的构建及意义  

Construction and significance of a vector for fruit-specific expression of porphyromonas gingivalis fimbriae fusion antigen gene

作　　者：戴海燕[1] 王华[2] 文少敏[1] 解娜[1] 

机构地区：[1]海南医学院口腔医学院,海口570102 [2]海南医学院海南省热带病重点实验室

出　　处：《山东医药》2011年第31期22-24,共3页Shandong Medical Journal

基　　金：海南省自然科学基金资助项目(807096)

摘　　要：目的构建含牙龈卟啉单胞菌菌毛融合抗原基因的番茄果实特异表达载体,并提高抗原基因的表达量和免疫原性,为其在番茄果实内表达和研制有效的转基因植物牙周炎疫苗奠定基础。方法通过PCR获得番茄果实特异表达启动子E8的核心序列片段(约1.11 kb),同时合成通过柔性接头连接的霍乱毒素B亚基(Cholera Tox-in B Subunit,CTB)和牙龈卟啉单胞菌菌毛FimA(266-337)(约600 bp)序列的基因片段,利用重叠区扩增基因拼接法将两基因片段连接,得到重组片段E8-CTB-FimA(266-337),双酶切E8-CTB-FimA(266-337)片段和植物表达载体pBI121,酶切产物连接后,获得重组载体pBI E8-CTB-FimA(266-337),利用酶切方法和测序方法对pBIE8-CTB-FimA(266-337)进行鉴定。结果基因测序及酶切结果表明特异表达载体已成功构建。结论本研究成功构建了含牙龈卟啉单胞菌菌毛融合抗原基因的番茄果实特异表达载体;该载体可用于转基因植物牙周炎疫苗的研制。Objective To construct a vector for tomato fruit-specific expression of perphyromonas 8~ngivalis fimbriac fusion antigen gene and to enhance the expression levels of the antigen gene as well as its immunogenicity, as the first step toward achieving its expression in tomato fruit and the development of an effective transgenic plant vaccine for periodontitis. Methods PCR was performed to collect the fragment （about 1.11 kb） of the tomato fruit-specific promoter ES. Mean-while, a recombinant fragment containing the cholera toxin B subunit （CTB） and FimA （266-337） （about 600 bp） genes was linked together with the use of a soft connector using gene splicing by overlap extension, resulting in a recombinant E8- CTB-FimA （266-337） fragment, which then together with the plant expression vector pBI121 underwent double restriction enzyme digestion. The recombinant vector pBIES-CTB-FimA （266-337） was obtained after joining the restriction enzyrae-digested fragments and was characterized by restriction enzyme digestion and sequencing methods. Results Results from gene sequencing and restriction enzyme digestion indicated successful construction of the recombinant plasmid. Conclusion In this study, a vector for tomato fruit-specific expression that contains the perphyromonas gingivalis fimbriac fusion antigen gene has been successfully constructed; the vector can be used in the development of a transgenic plant vaccine for periedontitis.

关 键 词：牙龈卟啉单胞菌 番茄果实特异启动子E8 菌毛蛋白 霍乱毒素B亚基 

分 类 号：R781.4[医药卫生—口腔医学]