In Vitro Screen of Bioinformatically Selected Bacillus anthracis Vaccine Candidates by Coupled Transcription, Translation, and Immunoprecipitation Analysis
The availability of the Bacillus anthracis genome sequence allowed for in silico selection of a few hundred open reading frames (ORFs) as putative vaccine candidates. To screen such a vast number of candidate ORFs, without resorting to laborious cloning and protein purification procedures, methods were developed for generation of PCR elements, compatible with in vitro transcription-translation and immunoprecipitation, as well as with their evaluation as DNA vaccines. Protocols will be provided for application of these methods to analyze the anti-B. anthracis antibody repertoire of hyperimmune sera or sera from convalescent and from DNA-vaccinated animals.
Key WordsIn vitro transcription in vitro translation reverse vaccinology vaccine development Bacillus anthracis bioinformatics geomics DNA vaccine gene gun immunoprecipitation serological analysis immunogenicity screen antigenicity screen
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