| 摘要: |
| 采用PCR方法扩增了吉林白鹅α干扰素（IFN-α）成熟肽编码序列，将IFN-α片段定向插入原核表达载体pGEX-6p-1中，构建重组质粒了pGEX-IFN-α，将重组质粒转入大肠杆菌BL21(DE3)的感受态细胞里，在IPTG诱导下表达可溶性的融合蛋白（GST-IFN-α）。SDS-PAGE、Western-blot检测结果表明，重组吉林白鹅IFN-α融合蛋白（rgIFN-α）的分子量大小约为43ku，表达量占菌体总蛋白的25%。重组吉林白鹅α干扰素经谷胱甘肽Sepharose-4B亲和柱层析纯化后，经过透析复性，得到纯化的目的蛋白，含量可达0.25mg/mL。将复性蛋白免疫新西兰白兔3次，制备高滴度的鹅IFN-α抗血清。本实验表达和纯化了鹅的IFN-α，并制备了兔抗鹅的IFN-α抗血清，为下一阶段关于鹅α干扰素的重组蛋白的应用奠定了基础。 |
| 关键词: 吉林白鹅 α干扰素 抗血清 |
| DOI： |
| 投稿时间：2011-12-22修订日期：2012-09-03 |
| 基金项目:吉林省科技发展计划项目（201101112）；吉林省教育厅“十二五”科学技术研究项目 |
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| Expression of JiLin White goose interferon-αin Escherichia coli and preparation of rabbit antisera against goose interferon-α |
| ligongmei |
| (College of Animal Science and Technology,JiLin Agricultural University) |
| Abstract: |
| αgene of JiLin White goose was amplified from genome DNA,the segment coding mature peptide of the GoIFN-αgene was cloned into prokaryotic expression vector pGEX-6p-1 to construct recombinant plasmid pGEX-IFN-α. pGEX-IFN-αwas transformed into E.coli BL-21(DE3),which can express fused protein GST-IFN-αin E.coli BL-21(DE3) by IPTG inducing. The expressed product was identified by SDS-PAGE and Western blot.The results showed that fusion protein GST-IFN-αwas about 45ku and it could react with interferon antibody.After purificating with Glutathione Sepharose-4B affinity column,and then refolded by dialysis against water.Antisera against goose IFN-αwas generated by immunizing rabbis with the purified protein.The expression and purification of recombinant goose IFN-αfusion protein and preparation of the antisera against goose IFN-αwill lay a foundation for the future application of goose IFN-α. |
| Key words: JiLin white goose IFN-α. gene antisera |
