| 摘要: |
| [目的]纯化Zhangfei融合蛋白并检测其免疫原性,为研究其在生殖内分泌调节中的作用提供理论依据.[方法]通过大肠杆菌原核表达系统进行Zhangfei融合蛋白的表达,筛选出最佳诱导表达条件,采用SDS-PAGE凝胶检测Zhangfei融合蛋白的表达情况.比较谷胱甘肽琼脂糖凝胶和电洗脱2种纯化Zhangfei融合蛋白的方法,并测定纯化的Zhangfei融合蛋白免疫Balb/c小鼠和家兔的效果.[结果]成功筛选出了表达Zhangfei融合蛋白的最佳诱导条件为IPTG终浓度3 mmol/L,诱导时间7 h;应用电洗脱方法可得到纯度较高的Zhangfei融合蛋白,其质量浓度为1.269 mg/mL,免疫小鼠的血清抗体效价达1 : 5×103以上,免疫兔的血清抗体效价高达1:3×106[结论]电洗脱纯化Zhangfei融合蛋白的方法优于谷胱甘肽琼脂糖凝胶纯化法,Zhangfei融合蛋白具有良好的免疫原性. |
| 关键词: Zhangfei融合蛋白 诱导表达 电洗脱 谷胱甘肽琼脂糖凝胶 抗体效价 |
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| 基金项目:国家自然科学基金项目（39970544）；西北农林科技大学青年骨干教师支持计划项目（01140305） |
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| Purification and immunifaction of Zhangfei Fusion Protein |
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| Abstract: |
| 【Objective】 The study purified Zhangfei Fusion Protein and detected the immunogenicity of Zhangfei Fusion Protein to offer theoretical basis for researching its regulations in geno-endocrine secretion.【Method】 Zhangfei Fusion Protein was expresed in the prokaryotic expression in E．coli to find the best way of expressing Zhangfei Fusion Protein.SDS-PAGE was used to detect Zhangfei Fusion Protein expression.The protein was separated by Glutathione Sepharose 4B and electroeluting and its immunifaction was detected.【Result】 The best way of expressing Zhangfei Fusion Protein in E．coli BL21:the final concentration of IPTG was 3 mmol/L,the time 7 h.Highly pure Zhangfei Fusion Protein was obtained by electroeluting.Valence of antibody in mouse and rabbit reached up to 1∶5×103 and 1∶3×106.【Conclusion】 Electroeluting to depurate Zhangfei Fusion Protein was better than Glutathione Sepharose 4B.Zhangfei Fusion Protein had a better immunogenicity. |
| Key words: Zhangfei Fusion Protein induced expression electroeluting Glutathione Sepharose 4B valence of antibody |
