葎草花粉主要变应原核酸疫苗pcDNA3.1-LC2的构建及其免疫原性鉴定

目的 构建葎草花粉主要变应原核酸疫苗pcDNA-LC2并鉴定其免疫原性.方法 将pTripIEx2-LC2质粒用EcoRⅠ和HindⅢ双酶切得到LC2葎草花粉变应原基因片段,将其插入pcDNA3.1(-)真核表达载体,在鉴定重组质粒构建成功后,大量制备去内毒素核酸疫苗pcDNA3.1-LC2,应用pcDNA3.1-LC2核酸疫苗免疫小鼠,取免疫血清行双向免疫扩散实验,分析其免疫原性.结果 通过测序确认构建pcDNA3.1-LC2中的672bp的编码框碱基与原序列一致,没有突变及缺失.应用pcDNA3.1-LC2免疫BALB/c小鼠,经双向免疫扩散试验验证pcDNA3.1-LC2在小鼠体内可以表达并产生特异性抗体.结论 成功的构建了葎草花粉主要变应原核酸疫苗pcDNA3.1-LC2,pcDNA3.1- LC2可在小鼠体内表达,并能产生葎草花粉变应原特异性抗体,具有良好的免疫原性.

Construction of humulus scandens pollen major allergen pcDNA3.1-LC2 DNA vaccine and study of its immunogenicity in mice

Objective To construct humulus scandens pollen major allergen DNA vaccine pcDNA-LC2 and evaluate its immunogenicity. Methods Humulus scandens pollen allergen gene was digested from recombinant plasmid pTripIEx2-LC2 by EcoRⅠ and HindⅢ of restriction endonuclease, and then inserted into expression vector pcDNA3.1(-). A large amount of endotoxin pcDNA3.1-LC2 purified plasmid was extracted after successful construction, and animal experiment was carried out to study its immunocompetence. Results Sequencing results showed that 672bp in the coding sequence of frames of recombinant pcDNA3.1-LC2 was in line with the original base line, without deletion or mutation. pcDNA3.1-LC2 of pollen allergen humulus scandens allergen vaccinated BALB/c mice. Double immunodiffusion test showed that pcDNA3.1-LC2 could be expressed and produce antibodies in mice. Conclusion Humulus scandens grass pollen major allergen DNA vaccine pcDNA3.1-LC2 has been successfully constructed. It can be expressed in mice, produces humulus scandens grass pollen allergen specific antibody and has a good immunogenicity.