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为研究DNA疫苗从细胞质到细胞核的过程,对免疫细胞中DNA疫苗的结合蛋白进行初步鉴定。提取小鼠脾脏免疫细胞的细胞质蛋白,将细胞质蛋白分别与DNA疫苗pVAX-OVA和空载体pVAX共孵育,孵育后首先由琼脂糖凝胶电泳分离与DNA结合的蛋白,然后通过SDS-PAGE进行分离和纯化,最后应用质谱技术分析其蛋白组分。质谱结果初步鉴定了免疫细胞中pVAX-OVA结合的蛋白有IQ motif containing F4等。免疫细胞中与pVAX结合的蛋白有Foxl2,SUV420H2和gamma actin等。Foxl2具有核定位序列,DNA结合区域和与核转运蛋白相互作用的特点,可能对DNA疫苗的进入具有促进作用。这些蛋白质是否可以影响DNA疫苗有效进入细胞核进行基因表达需要进一步研究。
In order to study the process of DNA vaccine from the cytoplasm to the nucleus, DNA vaccine binding proteins in immune cells were preliminarily identified. The cytoplasmic proteins of mouse spleen immune cells were extracted and the cytoplasmic proteins were co-incubated with DNA vaccine pVAX-OVA and empty vector pVAX respectively. After incubation, the DNA-binding proteins were separated by agarose gel electrophoresis and then separated by SDS-PAGE And purification, the final application of mass spectrometry analysis of its protein components. Mass spectrometry results preliminary identification of immune cells in the pVAX-OVA-binding protein IQ motif containing F4 and so on. The immune cells with pVAX binding protein Foxl2, SUV420H2 and gamma actin and so on. Foxl2 has a nuclear localization sequence, DNA binding region and the interaction with the nuclear transporter characteristics, DNA vaccine may have contributed to the entry. Whether these proteins can affect the DNA vaccine to effectively enter the nucleus for gene expression needs further study.