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目的 :在大肠杆菌中表达截短的YggG蛋白 ,并制备兔抗YggG截短体抗体。方法 :从含有大肠杆菌yggg基因全长DNA的质粒中 ,用PCR扩增截短的yggg基因序列 ,克隆入非融合表达载体pDH2中 ,构建YggG截短体的高表达工程菌 ,并温敏诱导表达非毒性的YggG蛋白 ,薄层扫描检测目的蛋白含量。免疫家兔制备兔抗YggG突变体抗血清 ,并以Western blot检测抗体效价、鉴定抗体特异性。结果 :大肠杆菌中表达的YggG截短突变体蛋白占菌体总蛋白的 3 1.7%。抗血清效价约 1∶2 0 0 0 ,Westernblot分析显示抗血清具有较高的特异性。结论 :成功地获得了YggG截短体蛋白 ,并制备了兔抗YggG突变体抗血清。用制备的该抗血清可检测到野生型全长YggG蛋白的表达
OBJECTIVE: To express truncated YggG protein in E. coli and prepare rabbit anti-YggG truncated antibody. Methods: The truncated yggg gene was amplified by PCR from the plasmid containing the full-length yggg gene of E. coli and cloned into the non-fusion expression vector pDH2 to construct a highly expressed engineering strain with YggG truncation. Expression of non-toxic YggG protein, TLC detection of the target protein content. Rabbits were immunized with rabbit anti-YggG mutant antiserum, and the antibody titers were detected by Western blot to identify the antibody specificity. Results: The YggG truncated mutant expressed in E. coli accounted for 31.7% of total bacterial proteins. Antiserum titer of about 1: 200, Western blot analysis showed that antisera with high specificity. Conclusion: YggG truncated protein was successfully obtained and rabbit anti-YggG mutant antiserum was prepared. The prepared wild-type full-length YggG protein was detected by using the prepared antiserum