DNA Recombination: Methods and Protocols

Part I. Genetic and Molecular Biological Approaches with Yeast

1. Methods to Study Mitotic Homologous Recombination and Genome Stability

Xiuzhong Zheng, Anastasiya Epstein, and Hannah L. Klein

2. Characterizing Resection at Random and Unique Chromosome Double-Strand Breaks and Telomere Ends

Wenjian Ma, Jim Westmoreland, Wataru Nakai, Anna Malkova, and Michael A. Resnick

3. Characterization of Meiotic Recombination Initiation Sites Using Pulsed-field Gel Electrophoresis

Sarah Farmer, Wing-Kit Leung, and Hideo Tsubouchi

4. Genome-Wide Detection of Meiotic DNA Double-Strand Break Hotspots Using Single-stranded DNA

Hannah G. Blitzblau and Andreas Hochwagen

5. Detection of Covalent DNA-Bound Spo11 and Topoisomerase Complexes

Edgar Hartsuiker

6. Molecular Assays to Investigate Chromatin Changes during DNA Double-Strand Break Repair in Yeast

Scott Houghtaling, Toyoko Tsukuda, and Mary Ann Osley

7. Analysis of Meiotic Recombination Intermediates by Two-Dimensional Gel Electrophoresis

Jasvinder A. Singh and G. Valentin Börner

8. Mapping of Crossover Sites Using DNA Microarrays

Stacy Y. Chen and Jennifer C. Fung

9. Using the Semi-Synthetic Epitope System to Identify Direct Substrates of the Meiosis-Specific Budding Yeast Kinase, Mek1

Hsiao-Chi Lo and Nancy M. Hollingsworth

10. Genetic and Molecular Analysis of Mitotic Recombination in Saccharomyces cerevisiae

Belén Gómez-González, José F. Ruiz, and Andrés Aguilera

11. In vivo Site-Specific Mutagenesis and Gene Collage Using the Delitto Perfetto System in Yeast Saccharomyces cerevisiae

Samantha Stuckey, Kuntal Mukherjee, and Francesca Storici

12. Detection of RNA-Templated Double-Strand Break Repair in Yeast

Ying Shen and Francesca Storici

Part II. Genetic and Molecular Biological Approaches with Higher Eukaryotes

13. SNP-Based Mapping of Crossover Recombination in C. elegans

Grace C. Bazan and Kenneth J. Hillers

14. Characterization of Meiotic Crossovers in Pollen from Arabidopsis thaliana

Jan Drouaud and Christine Mézard

15. Isolation of Meiotic Recombinants from Mouse Sperm

Francesca Cole and Maria Jasin

16. Homologous Recombination Assay for Interstrand Crosslink Repair

Koji Nakanishi, Francesca Cavallo, Erika Brunet, and Maria Jasin

17. Evaluation of Homologous Recombinational Repair in Chicken B Lymphoma Cell Line, DT40

Hiroyuki Kitao, Seiki Hirano, and Minoru Takata

18. Understanding the Immunoglobulin Locus Specificity of Hypermutation

Vera Batrak, Artem Blagodatski, and Jean-Marie Buerstedde

Part III. In Vitro Reconstitution of Homologous Recombination Reactions and Single Molecular Analysis of Recombination Proteins

19. Quality Control of Purified Proteins Involved in Homologous Recombination

Xiao-Ping Zhang and Wolf-Dietrich Heyer

20. Assays for Structure-Selective DNA Endonucleases

William D. Wright, Kirk T. Ehmsen, and Wolf-Dietrich Heyer

21. In vitro Assays for DNA Pairing and Recombination-Associated DNA Synthesis

Jie Liu, Jessica Sneeden, and Wolf-Dietrich Heyer

22. An In vitro Assay for Monitoring the Formation and Branch Migration of Holliday Junctions Mediated by a Eukaryotic Recombinase

Yasuto Murayama and Hiroshi Iwasaki

23. Reconstituting the Key Steps of the DNA Double-Strand Break Repair In vitro

Matth

Homologous recombination is important in various aspects of DNA metabolism, including damage repair, replication, telomere maintenance, and meiosis, and yeast genetics has successfully provided a framework for the mechanism of homologous recombination. Divided into four convenient sections, DNA Recombination: Methods and Protocols covers recent techniques that best utilize the advantages of the yeast system, prescribing to the belief that yeast will keep serving as a great model organism to study homologous recombination. Chapters have also been included for such exceptions as the group of genes involved in recombination that are found solely in higher eukaryotes, such as BRCA2. And looking forward, a necessary step in the direction of understanding the homologous recombination process is to isolate the machine and let it work in a test tube. Understanding the design by studying the appearance and behavior of the machinery as a single molecule will be an important milestone toward understanding the mechanism of action of the machinery. Techniques covering these topics have also been included. Written in the 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 protocols, and notes on troubleshooting and avoiding known pitfalls.

Authoritative and easily accessible, DNA Recombination: Methods and Protocols serves as an ideal guide to scientists of all backgrounds with its well-honed methodologies and strives to bring the reader to the next level of understanding regarding this vital subject.