Differential expression of two isolates of beak and feather disease virus capsid protein in Escherichia coli

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• 作者：Edward I. Patterson^a ; ^c ; Crystall M.D. Swarbrick^b ; ^c ; Noelia Roman^b ; Jade K. Forwood^b ; ^c ; Shane R. Raidal^a ; ^c ; ^{shraidal@csu.edu.au}
• 关键词：Beak and feather disease virus ; BFDV ; Capsid-associated protein ; Protein purification
• 刊名：Journal of Virological Methods
• 出版年：2013
• 出版时间：April, 2013
• 年：2013
• 卷：189
• 期：1
• 页码：118-124
• 全文大小：868 K

文摘

Expression of recombinant beak and feather disease virus (BFDV) capsid-associated protein (Cap) has relied on inefficient techniques that typically produce low yields or use specialized expression systems, which greatly increase the cost and expertise required for mass production. An Escherichia coli system was used to express recombinant BFDV Cap derived from two isolates of BFDV, from a Long-billed Corella (Cacatua tenuirostris) and an Orange-bellied parrot (OBP; Neophema chrysogaster). Purification by affinity and size exclusion chromatography was optimized through an iterative process involving screening and modification of buffer constituents and pH. A buffer containing glycerol, ¦Â-mercaptoethanol, Triton X-100, and a high concentration of NaCl at pH 8 was used to increase solubility of the protein. The final concentration of the corella-isolated BFDV protein was fifteen- to twenty-fold greater than that produced in previous publications using E. coli expression systems. Immunoassays were used to confirm the specific antigenicity of recombinant Cap, verifying its validity for use in continued experimentation as a potential vaccine, a reagent in diagnostic assays, and as a concentrated sample for biological discoveries.