The field of eukaryotic DNA repair is enjoying a period of remarkable growth and discovery, fueled by technological advances in molecular bi- ogy, protein biochemistry, and genetics. Notable achievements include the molecular cloning of multiple genes associated with classical human repair disorders, such as xeroderma pigmentosum, Cockayne syndrome, and ataxia telangiectasia; elucidation of the core reaction of nucleotide excision repair (NER); the discovery that certain NER proteins participate not only in repair, but also in transcription; recognition of the crucial role played by mismatch repair processes in maintenance of genome stability and avoidance of cancer; the findings that the tumor suppressor protein p53 is mutated in many types of cancer, and has a key role in directing potentially malignant, genotoxin-d- aged cells towards an apoptotic fate; and the discovery and elaboration of DNA damage (and replication) checkpoints, which placed repair phenomen- ogy firmly within a cell-cycle context. Of course, much remains to be learned about DNA repair. To that end, DNA Repair Protocols: Eukaryotic Systems is about the tools and techniques that have helped propel the DNA repair field into the mainstream of biological research. DNA Repair Protocols: Eukaryotic Systems provides detailed, step-- step instructions for studying manifold aspects of the eukaryotic response to genomic injury. The majority of chapters describe methods for analyzing DNA repair processes in mammalian cells. However, many of those techniques can be applied with only minor modification to other systems, and vice versa.

Part I. Mutant Isolation and Gene Cloning

Isolation of DNA Structure-Dependent Checkpoint Mutants in S. pombe
Rui G. Martinho and Antony M. Carr

Isolating Mutants of the Nematode Caenorhabditis elegans That Are Hypersensitive to DNA-Damaging Agents
Phil S. Hartman and Naoaki Ishii

Isolating DNA Repair Mutants of Drosophila melanogaster, Daryl S. Henderson

Generation, Identification, and Characterization of Repair-Defective Mutants of Arabidopsis
Anne Britt and Cai-Zhong Jiang

Screening for g-Ray Hypersensitive Mutants of Arabidopsis
Corinne S. Davies

Isolation of Mutagen-Sensitive Chinese Hamster Cell Lines by Replica Plating
Malgorzata Z. Zdzienicka

Strategies for Cloning Mammalian DNA Repair Genes
Larry H. Thompson

Novel Complementation Assays for DNA Repair-Deficient Cells: Transient and Stable Expression of DNA Repair Genes
Lin Zeng, Alain Sarasin, and Mauro Mezzina

Part II. Recognition and Removal of Inappropriate or Damaged DNA Bases

The Use of Electrophoretic Mobility Shift Assays to Study DNA Repair
Byung Joon Hwang,Vaughn Smider, and Gilbert Chu

Mismatch Repair Assay
Stephanie E. Corrette-Bennett and Robert S. Lahue

Measurement of Activities of Cyclobutane-Pyrimidine-Dimer and (6-4)-Photoproduct Photolyases
John B. Hays and Peter Hoffman

A Dot Blot Immunoassay for UV Photoproducts
Shirley McCready

Measurement of UV Radiation-Induced DNA Damage Using Specific Antibodies
Ann E. Stapleton

Quantification of DNA Photoproducts in Mammalian Cell DNA Using Radioimmunoassay
David L. Mitchell

Monitoring Removal of Cyclobutane Pyrimidine Dimers in Arabidopsis
John B. Hays and Qishen Pang

DNA Damage Quantitation by Alkaline Gel Electrophoresis
Betsy M. Sutherland, Paula V. Bennett, and John C. Sutherland

The Comet Assay (Single-Cell Gel Test): A Sensitive Genotoxicity Test for the Detection of DNA Damage and Repair
Günter Speit and Andreas Hartmann

Measuring the Formation and Repair of UV Photoproducts by Ligation-Mediated PCR
Gerd P. Pfeifer and Reinhard Dammann

PCR-Based Assays for Strand-Specific Measurement of DNA Damage and Repair I: Strand-Specific Quantitative PCR
Keith A. Grimaldi, John P. Bingham, and John A. Hartley

PCR-Based Assays for Strand-Specific Measurement of DNA Damage and Repair II: Single-Strand Ligation-PCR
Keith A. Grimaldi, Simon R. McAdam, and John A. Hartley

Gene-Specific and Mitochondrial Repair of Oxidative DNA Damage
R. Michael Anson and Vilhelm A. Bohr

Characterization of DNA Strand Cleavage by Enzymes That Act at Abasic Sites in DNA
Walter A. Deutsch and Adly Yacoub

Base Excision Repair Assay Using Xenopus laevis Oocyte Extracts
Yoshihiro Matsumoto

In Vitro Base Excision Repair Assay Using Mammalian Cell Extracts
Guido Frosina, Enrico Cappelli, Paola Fortini, and Eugenia Dogliotti

Nucleotide Excision Repair in Saccharomyces cerevisiae Whole-Cell Extracts
Johnson M. S. Wong, Zhigang He, and C. James Ingles

In Vitro Excision Repair Assay in Schizosaccharomyces pombe
Bernard Salles and Patrick Calsou

Nucleotide Excision Repair Assay in Drosophila melanogaster Using Established Cell Lines
Kenji Kohno and Takuya Shimamoto

Nucleotide Excision Repair in Nuclear Extracts from Xenopus Oocytes
Eric J. Ackerman, Lilia K. Koriazova, Jitendra K. Saxena, and Alexander Y. Spoonde

Assay for Nucleotide Excision Repair Protein Activity Using Fractionated Cell Extracts and UV-Damaged Plasmid DNA
Maureen Biggerstaff and Richard D. Wood

Dual-Incision Assays for Nucleotide Excision Repair Using DNA with a Lesion at a Specific Site
Mahmud K.