pcD-awte候选疟疾DNA疫苗制备及其质量相关性分析

本实验室构建的疟疾DNA疫苗经动物试验表明具有很好的免疫原性，为申请临床试验，进行了制备工艺的研究。本研究将含pcD-awte质粒的大肠杆菌DH5α在发酵罐中发酵培养，碱裂解法粗提质粒，再依次通过Sepharose 6FF分子筛层析、Plasmidselect 亲硫吸附层析和Source 30Q离子交换层析精制获得质粒纯品，并对纯品进行质量分析。结果每升培养液可获得质粒纯品43.9mg，质量符合Ferreira等推荐的药用标准。

Preparation and Quality Assay of pcD-awte Candidate Malaria DNA Vaccine

The pcD-awte candidate malaria DNA vaccine could induce protective immune-responses through animal test, so it is necessary to establish a suitable preparative protocol with which the vaccine can be applied in clinical test as soon as possible. Here we reported an efficient preparative method, with which plasmid DNA vaccine could be produced in high yield and purity. Escherichia coli DH5α containing recombinant plasmid pcD-awte was grown through fed-batch fermentation in 5L fermentor with basic culture media of improved TB. After fermentation, the crude plasmid DNA　was obtained by alkaline lysis and the plasmid DNA was further purified with a three-step efficient chromatographic procedure, including Sepharose 6FF size exclusion chromatography , Plasmidselect thiophilic aromatic chromatography and Source 30Q anion exchange chromatography. The purified plasmid DNA was identified, which showed that the yield of final product reached 43.9 mg/L and that the quality met the standards of pharmaceutical-grade plasmid DNA.