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This volume explores homologous recombination's (HR) essential role in meiotic and somatic cells. It discusses the analysis of different steps of the HR process from the genetic, molecular biology, and cell biology perspectives. Using a variety of model systems, chapters in this book cover topics such as the genome-wide mapping of DNA double-strand breaks (DSB); analysis of DNA-end resection and recombination intermediates by gel electrophoresis and southern blotting; cell-based monitoring of HR activity in meiotic and mitotic cells; in vitro analysis of branch migration, DNA strand exchange and Holliday junction resolution; or super resolution imaging of HR repair at collapsed replication forks. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.
Cutting-edge and thorough, Homologous Recombination: Methods and Protocols is a valuable resource that uses both classical and more recent approaches to answer questions on the HR mechanism. It is a useful tool for scientists working on the field of genome integrity, as well as those working in cancer biology and DNA repair.
List of contents
Detection of DNA Double Strand Breaks by Gamma-H2AX Immunodetection.- END-seq: An Unbiased, High-Resolution and Genome-Wide Approach to Map DNA Double Strand Breaks and Resection in Human Cells.- Resection of a DNA Double-Strand Break by Alkaline Gel Electrophoresis and Southern Blotting.- Analysis of DNA Double-Strand Break Ends Resection and Single-Strand Annealing in S. pombe.- Quantifying DNA End Resection in Human Cells.- Genetic and Molecular approaches to Study Chromosomal Breakage at Secondary Structure-Forming Repeats.- Biochemical analysis of D-Loop Extension and DNA Strand Displacement Synthesis.- DNA Strand Exchange to Monitor Human RAD51-Mediated Strand Invasion and Pairing.- Monitoring Homologous Recombination Activity in Human Cells.- Inter-Homolog Homologous Recombination in Mouse Embryonic Stem Cells.- Branch Migration Activity of Rad54 Protein.- Holliday Junction Resolution.- Identification and Analysis of Different Types of UFBs.- Intrachromosomal Recombination in Yeast.- Genome-Wide Analysis of Mitotic Recombination in Budding Yeast.- Monitoring Gene Conversion in Budding Yeast by Southern Blot Analysis.- DNA Double-Strand Break Induced Gene Amplification in Yeast.- Measuring Chromosome Pairing during Homologous Recombination in Yeast.- Cytological Monitoring of Meiotic Crossovers in Spermatocytes and Oocytes.- Detection of DSBs in C. elegans Meiosis.- Methods to Map Meiotic Recombination Proteins in Saccharomyces cerevisiae.- Investigation of Break-Induced Replication in Yeast.- Measurement of Homologous Recombination at Stalled Mammalian Replication Forks.- Super Resolution Imaging of Homologous Recombination Repair at Collapsed Replication Forks.- The Analysis of Recombination-Dependent Processing of Blocked Replication Forks by Bi-Dimensional Gel Electrophoresis.- The Sister Chromatid Exchange Assay in Human Cells.- Analysis of Recombination at Yeast Telomeres.- Gel Electrophoresis Analysis of rDNA Instability in Saccharomyces cerevisiae.- Analyzing Homologous Recombination at a Genome Wide Level.- CRISPR/Cas9 Induced Breaks in Heterochromatin, Visualized by Immunofluorescence.- In Vivo Binding of Recombination Proteins of Non-DSB DNA Lesions to Replication Forks.- Live Cell Imaging of Nuclear Actin Filaments and Heterochromatic Repair foci in Drosophila and Mouse Cells.- In Vitro Characterization of Sumoylation of HR Proteins.- High-Throughput Analysis of Heteroduplex DNA in Mitotic Recombination Products.- Fluorescence Microscopy for Analysis of Relocalization of Structure-Specific Endonucleases.- Physical and Genetic Assays for the Study of DNA Joint Molecules Metabolism and Multi-Invasion-Induced Rearrangements in S. cerevisiae.
