IL-15基因真核表达质粒的构建及其对CVB3 VP1基因疫苗的免疫增强作用

CONSTRUCTION OF EUKARYOTIC EXPRESSION PLASMID pcDNA3-IL-15 AND ITS IMMUNOLOGICAL ENHANCEMENT EFFECT TO CVB3 VP1 GENE VACCINE

目的探讨人白细胞介素 15 (humaninterleukin 15 ,hIL 15 )基因佐剂对柯萨奇B3病毒 (CoxsackievirusB3 ,CVB3 )VP1基因疫苗的免疫增强效果。方法从人胚肺二倍体细胞提取hIL 15的mRNA ,用逆转录 聚合酶链反应 (reversetranscriptase polymerasechainreaction ,RT PCR)扩增出cDNA ,构建成真核表达克隆 pcDNA3 IL 15。将pcDNA3 VP1、pcDNA3 VP1+ pcDNA3 IL 15、pcDNA3 IL 15、pcDNA3和盐水分别肌内注射Balb/c小鼠 ,每次免疫后 6天取血清用微量中和试验测CVB3中和抗体 ,3次免疫后用 80 0TCID5 0CVB3感染小鼠 ,观察小鼠的生存率。结果 pcDNA3 IL 15重组质粒的目的基因与hIL 15基因序列相同 ;pcDNA3 VP1和pcDNA3 VP1+ pcDNA3 IL 15能诱导小鼠产生中和抗体 ,抗体滴度随免疫次数增加而提高 ;pcDNA3 VP1+ pcDNA3 IL 15组抗体滴度和生存率均高于 pcDNA3 VP1组。结论本研究成功构建了hIL 15重组质粒 ,该质粒对真核表达重组质粒 pcDNA3

ObjectiveTo study the immunological enhancing effect of human interleukin 15(hIL 15) as gene adjuvant on Coxsackievirus B3(CVB3) VP1 gene vaccine.MethodsThe mRNA of IL 15 was extracted from embryonic pulmonary diploid cells,then amplified with reverse transcriptase polymerase chain reaction(RT PCR) and inserted into plasmid pcDNA3 to construct a eukaryotic expression plasmid pcDNA3 IL 15. Balb/c mice were divided into pcDNA3 VP1?pcDNA3 VP1+pcDNA3 IL 15?pcDNA3 IL 15? pcDNA3 and saline control groups, and injected with corresponding agent respectively in quadrideps. Sera were collected in the sixth day after every immunization for measuring neutralizing antibodies.The mice were infected with 800TCID50 CVB3 after three times of immunization injection and the survival was observed.ResultsThe sequence of inserted gene fragment of pcDNA3 IL 15 was identical to that of hIL 15. pcDNA3 VP1 and pcDNA3 VP1+pcDNA3 IL 15 could induce the production of neutralizing antibodies and the titers of antibody increased with the times of immunization. In pcDNA3 VP1+pcDNA3 IL 15 group the antibody titers and survival were higher than that of pcDNA3 VP1 group. Conclusion pcDNA3 IL 15 was constructed successfully and it could enhance the immunological effect of pcDNA3 VP1 vaccine.