Trinucleotide Repeat Protocols

Established leaders in trinucleotide repeat disease describe in step-by-step detail their best techniques for studying trinucleotide pathology at the molecular level. The protocols cover a variety of targets, ranging from DNA and RNA to proteins and whole animals, and focus not only on causal genes, but also on their consequent affected products,

Includes supplementary material: sn.pub/extras

Klappentext
The discovery that trinucleotide repeats significantly influence the age of onset and severity of a variety of hereditary neurological disorders has opened the door to a deeper understanding of the disease mechanisms involved, as well as to a more productive search for novel therapeutic interventions. In Trinucleotide Repeat Protocols, established leaders in trinucleotide repeat disease describe in step-by-step detail their best techniques for studying trinucleotide pathology at the molecular level. The protocols cover a variety of targets, ranging from DNA and RNA to proteins and whole animals, and focus not only on causal genes, but also on their consequent products, such as transcription factors, neurotransmitter receptors, proteasomes, and mitochondria/oxidation damage. Experimental systems employed include E. coli, yeast, C. elegans, mouse, and generally take a clinical point of view. The authors utilize a wide range of techniques, including gel electrophoresis, quantitative RT-PCR, immunological analysis, antibody usage and its applications, receptor assays using radioisotope handling, gene delivery by virus, brain cell and organotypic cultures, gender dependency, and neuron structure analysis. Each protocol follows the successful Methods in Molecular Biology™ series format, offering step-by-step laboratory instructions, an introduction outlining the principle behind the technique, lists of the necessary equipment and reagents, and tips on troubleshooting and avoiding known pitfalls.
Cutting-edge and highly practical, Trinucleotide Repeat Protocols offers neuroscientists powerful tools to elucidate both normal brain function and the mechanisms of hereditary neurological disease, as well as to develop the next generation of therapies for neuronal genetic diseases.

Inhalt
Mouse Models of Triplet Repeat Diseases.- Analysis of Triplet Pepeat DNAs and RNAs.- Analysis of Triplet Repeat Replication by Two-Dimensional Gel Electrophoresis.- Genetic Assays for Triplet Repeat Instability in Yeast.- Detection and Isolation of Trinucleotide Repeat Expansions Using the RED Method.- Analysis of Unstable Triplet Repeats Using Small-Pool Polymerase Chain Reaction.- Real-Time RT-PCR for CTG Repeat-Containing Genes.- Detection and Analysis of Polyglutamine-Containing Proteins and Their Aggregates.- Antibodies Against Huntingtin.- Using Antibodies to Analyze Polyglutamine Stretches.- Solubilization of Aggregates Formed by Expanded Polyglutamine Tract Expression in Cultured Cells.- Establishment of Animal and Cultured Cell Models for Trinucleotide Repeat Diseases.- Caenorhabditis elegans as a Model System for Triplet Repeat Diseases.- Monitoring Aggregate Formation in Organotypic Slice Cultures From Transgenic Mice.- The CGG Repeat and the FMR1 Gene.- Analysis of CTG Repeats Using DM1 Model Mice.- Lentiviral-Mediated Gene Transfer to Model Triplet Repeat Disorders.- Mouse Tissue Culture Models of Unstable Triplet Repeats.- In Vivo Analysis of Trinucleotide Repeat Diseases.- Neurotransmitter Receptor Analysis in Transgenic Mouse Models.- Chromatin Immunoprecipitation Technique for Study of Transcriptional Dysregulation in Intact Mouse Brain.- Techniques for Thick-Section Golgi Impregnation of Formalin-Fixed Brain Tissue.- Assessment of Impaired Proteasomal Function in a Cellular Model of Polyglutamine Diseases.- Assessment of In Vitro and In Vivo Mitochondrial Function in Friedreich's Ataxia and Huntington's Disease.- Triplet Repeats and DNA Repair.- Oxidative Damage in Huntington's Disease.