| 噬菌体Bp7尾丝蛋白gp37的原核表达与鉴定 |
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| 引用本文： | 孙虎芝,任慧英,张灿.噬菌体Bp7尾丝蛋白gp37的原核表达与鉴定[J].动物医学进展,2016(5):10-14. |
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| 作者姓名： | 孙虎芝 任慧英 张灿 |
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| 作者单位： | 青岛农业大学动物科技学院,山东青岛,266109 |
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| 基金项目： | 青岛农业大学创新计划项目(QYC201406)；山东省自然科学基金项目(ZR2013CQ024；ZR2015CM020) |
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| 摘 要： | 为了研究噬菌体尾丝蛋白gp37在识别宿主菌大肠埃希菌K12过程中的作用,PCR扩增获得gp37基因C端1 161bp序列,以EGFP为荧光标签,构建重组表达质粒pET-28a-EGFP-gp37,在大肠埃希菌BL21(DE3)中IPTG诱导表达重组蛋白EGFP-gp37,亲和层析纯化蛋白。SDS-PAGE及Western blot检测结果表明,重组蛋白EGFP-gp37在大肠埃希菌BL21(DE3)为可溶性表达,表达量占菌体总蛋白量的23%。EGFP-gp37融合蛋白与大肠埃希菌K12相互作用,激光共聚焦显微镜检测结果显示,尾丝蛋白gp37能够吸附到宿主菌K12表面,表明尾丝蛋白gp37参与宿主菌受体的识别和吸附过程。
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| 关 键 词： | 噬菌体Bp7 尾丝蛋白gp37 绿色荧光蛋白EGFP 大肠埃希菌K12 |
Prodaryotic Expression and Activity Determination of Tail Fiber Protein gp37 of Phage Bp7 |
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| Abstract: | In order to identify the tail fiber protein gp37 function in phage Bp7 adsorbing on E.coli K12, 1 161 bp C terminal sequence of gp37 as amplified by PCR,and recombinant plasmid pET-28a-EGFP-gp37 was constructed with a fluorescent tag of EGFP.In E.coli BL21(DE3),the fusion protein EGFP-gp37 was induced by IPTG to express and purified by affinity chromatography.SDS-PAGE and Western blot analysis showed that protein EGFP-gp37 was expressed in supernatant of E.coli BL21(DE3).The expression level reached 23%.The purfied protein of EGFP-gp37 was incubated with E.coli K12 and detected by laser con-focal microscopy.The result showed that protein EGFP-gp37 adsorbed on the surface of E.coli K12,which indicated that tail fiber protein gp37 participated in the receptor recognition of phage Bp7. |
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| Keywords: | phage Bp7 tail fiber protein gp37 green fluorescent protein EGFP E coli K12 |
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