摘要
为了研究赖型钩体内鞭毛蛋白基因在动物体内的表达及其免疫保护性，采用ＰＣＲ扩增内鞭毛蛋白编码基因，以表达质粒ＶＲ１０１２为载体，构建了含有钩体的内鞭毛基因的重组质粒。在新西兰大白兔四头肌内注射该重组质粒ＤＮＡ５００μｇ／只，重复３次，每次间隔３周，于每次注射前和末次注射后３周，检测血清抗赖型钩体抗体效价。结果显示：在第１次加强免疫注射后３周，试验组血清抗体效价显著增高。豚鼠免疫保护试验表明，该ＤＮＡ重组质粒有明显免疫保护作用，在无佐剂条件下试验组存活率为９０％（９／１０只）。本研究为钩体核酸疫苗研制奠定了基础。
Leptospirosis is one of the most widespread zoonosis. Immunization of NZW rabbit with
plasmid DNA constructs encoding leptospiral endoflagellin can induce specific antibody
response in the animal. The vector, VR1012 consists of a pUC18 backbone with the
cytomegalovirus(CMV) IE1 enhancer, promoter, and intron A transcription regulatory elements
and the BGH polyadenylation sequences driving the expression of leptospiral endoflagellar
gene,flaB2. In this study, a pair of oligonucleotide primers were designed to amplify the
endoflagellar of L.interrogans serovar lai. An approximately 840bp fragment was generated with
PCR and inserted into expressed plasmid vector, VR1012, after the fragment of VR1012 were
digested respectively with EcoRV and Sal. A recombinant plasmid (designated VR1012+flaB2)
was obtained. Plasmid encoding leptospiral endoflagellin was injected into quadriceps of NZW
rabbits.This resulted in the generation of specific leptospiral antibody with very high ELISA titer.
Immunoprotection was performed in guinea pigs. The group VR1012+flaB2 had higher survival
rate, compared with the group VR1012 only (control) and the group Normal Saline
only(control).The result suggests that the technique of DNA injection should have an advantage
over certain previous vaccine technologies.
出处
《华西医科大学学报》
CSCD
1999年第2期128-132,共5页
Journal of West China University of Medical Sciences
基金
高等学校博士学科点专项基金
卫生部和四川省科委科研基金
关键词
钩端螺旋体
DNA疫苗
免疫原性
免疫保护
LeptospiraDNA recombinantInjections
intramuscularEndoflagellinDNA vaccinesImmunoprotection
