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This protocol book includes common and advanced research protocols for Yersinia pestis research and for its identification, genotyping and source-tracing. This book includes protocols for genomic and transcriptomic analysis, small RNA study, protein-protein interaction, gene deletion. It will be helpful for graduate students, clinicians, and researchers in infectious diseases, systems biology, and evolution and numerous other relevant fields.
Sommario
Isolation and identification of Yersinia pestis.- Isolation and characterization of Yersinia pestis bacteriophages.- Extraction of chromosomal DNA and plasmid from Yersinia pestis.- Extraction of total RNAs from Yersinia pestis.- Genotyping of Yersinia pestis.- Genome-wide variation analysis of Yersinia pestis.- Genome-wide detection of expressed sRNAs in Y. pestis with RNA-seq.- Validation of Y. pestis sRNAs with nonradiative-labeled Northern Blot.- Determination of transcriptional start sites (TSSs) in Y. pestis with a primer extension assay.- Purification of Hfq-associated RNAs with RNA immunoprecipitation (RIP).- Overexpression of target sRNA in tightly controlled multicopy plasmids.- Target validation of sRNA with a GFP reporter gene fusion system.- Inducible gene expression in Yersinia pestis.- Analysis of Yops secretion by Y. pestis Type III secretion system.- Co-Immunoprecipitation analysisfor detection of protein-protein interaction in Y. pestis.- Detection of type III secretion system needle assembly in Yersinia pestis by cross-linking of YscF.- Evaluation of protein translocation into eukaryotic cells by Y. pestis.- Adhesion analysis of Y. pestis to the host cells.- Construction of gene deletion mutants in Yersinia pestis.- CRISPR-Cas12a-assisted recombineering in Yersinia pestis.- Plasmid curing in Yersinia pestis.- Genetic regulation analysis of Yersinia pestis.- Investigation of Biofilm formation in Y. pestis.- Extraction and purification of F1 capsule antigen from Y. pestis.- Screening of regulator responses to Yersinia pestis infection with a concatenated tandem array of transcription factor response elements-pull down.- Animal models for Yersinia pestis.- Antibiotic resistance evaluation of Yersinia pestis.- PCR for detection and identification of Yersinia pestis.- Immunological methods for detection and identification of Yersinia pestis.- Analysis of fatty acids in Yersinia pestis by gas chromatography.- Evaluation of efficacy of Yersinia pestis vaccine.- Online resources for Yersinia pestis.- Effects of Y. pestis on macrophage.
Info autore
Dr. Ruifu Yang is a professor at the Beijing Institute of Microbiology and Epidemiology and the Chief-Scientist for National Basic Research Program (973 Program). He serves as the vice president for the Chinese Society for Microbiology. His research focuses on bacterial pathogenesis using genomics- and proteomics-based techniques. He has published more than 100 articles on international well-known journals, such as Nature Genetics, PNAS, BMC Microbiology.
Riassunto
This protocol book includes common and advanced research protocols for Yersinia pestis research and for its identification, genotyping and source-tracing. This book includes protocols for genomic and transcriptomic analysis, small RNA study, protein-protein interaction, gene deletion. It will be helpful for graduate students, clinicians, and researchers in infectious diseases, systems biology, and evolution and numerous other relevant fields.
