Full Description
The discovery of Epstein-Barr virus (EBV) by Epstein, Achong, and Barr, reported in 1964 (Lancet 1:702-703), was stimulated by Denis Burkitt's rec- nition of a novel African childhood lymphoma and his postulation that an infectious agent was involved in the tumor's etiology (Nature194:232-234, 1962). Since then, molecular and cellular biological and computational technologies have progressed by leaps and bounds. The advent of recombinant DNA technology opened the possibilities of genetic research more than most would have realized. Not only have the molecular tools permitted the analyses of viral mechanisms, but, importantly, they have formed the basis for discerning viral presence and, subsequently, viral involvement in an increasing number of diseases. Though in every field of science the search for further knowledge is likely to be a limitless phenomenon, the distinct goal in EBV research, namely, to gain sufficient insight into the viral-host interaction to be able to intercept the pathogenic process, is beginning to be realized. Epstein-Barr virus research has effectively entered the postgenomic era that began with the sequencing of the first strains, cloned in the mid to late 1980s.
Contents
Genome and Transcript Analyses.- Epstein-Barr Virus.- Analysis of Replication of oriP-Based Plasmids by Quantitative, Competitive PCR.- Genetic Analysis and Gene Expression with Mini-Epstein-Barr Virus Plasmids.- Construction of cDNA Libraries for the Analysis of the Structure of Complementary Strand Transcripts (CSTs).- Analysis of the Expression and Function of the EBV-Encoded Small RNAs, the EBERs, in Heterologous Cells.- Visualizing EBV Expression Patterns by FISH.- Viral Detection.- In Situ Detection of Epstein-Barr Virus DNA and Viral Gene Products.- Phenotype Determination of Epstein-Barr Virus-Infected Cells in Tissue Sections.- Detection of EBV Infection at the Single-Cell Level.- Detection and Discrimination of Latent and Replicative Herpesvirus Infection at the Single Cell Level In Vivo.- Culture Methods.- Virus Isolation.- Generation of Lymphoblastoid Cell Lines (LCLs).- Cell Cycle Distribution of B-Lymphocytes and Cell Lines.- of Plasmid Vectors into Cells Via Electroporation.- Malignant Transformation and Immortalization Assays in Animal Cells Transfected with the BARF1 Gene.- Transient Gene Expression and MACS Enrichment.- In Vitro Assays to Study Epithelial Cell Growth.- In Vitro Assays to Study Epithelial Cell Differentiation.- Cell Sensitivity Assays.- Qualitative Detection of Apoptotic Cells Assessed by DNA Fragmentation.- Immune Assays.- Regression Assay.- Generation of Polyclonal EBV-Specific CTL Cultures and Clones.- Determination of Antigen and Fine Peptide Specificity of EBV-Specific CTLs.- Limiting Dilution Assay.- Viral Protein Detection.- Antibodies for Detecting EBV Latent Proteins.- Detection of EBV Latent Proteins by Western Blotting.- Biosynthetic Radiolabeling of Virus Glycoproteins for Immunoprecipitation and Electrophoretic Analysis.- Protein-Protein, Protein-DNA, and Protein-RNA Interactions.- The Yeast Two-Hybrid Assay to Identify Interacting Proteins.- Identification of Transactivation, Repression, and Protein-Protein Interaction Domains Using GAL4-Fusion Proteins.- Magnetic DNA Affinity Purification of a Cellular Transcription Factor.- Screening an Expression Library with a DNA Binding Site.- Analysis of DNA Binding Proteins by Mobility Shift Assay.- Analysis of RNA-Protein Interactions of the EBV-Encoded Small RNAs, the EBERs.- Protein Activity Assays.- Chimeric and Mutated Variants of LMP1.- Assaying the Activity of Kinases Regulated by LMP1.- In Vitro Assays for the Detection of Protein Tyrosine Phosphorylation and Protein Tyrosine Kinase Activities.- Tissue and In Vivo Protocols.- Analysis of Apoptosis in Tissue Sections.- Considerations in Generating Transgenic Mice.- In Vivo Assay of Cellular Proliferation.- Topical Chemical Carcinogen Treatment in Mice.- Separation of Epidermal Tissue from Underlying Dermis and Primary Keratinocyte Culture.- Selection and Enrichment of B Cells from Lymphoid Tissues.- Detection of Immunoglobulin Gene Rearrangements.
