Genetic and Molecular Approaches to Study Chromosomal Breakage at Secondary Structure–Forming Repeats
| Autor: | Anissia Ait Saada, Kirill S. Lobachev, Alex B Costa |
|---|---|
| Rok vydání: | 2020 |
| Předmět: |
Genome instability
Inverted repeat Saccharomyces cerevisiae Computational biology Article 03 medical and health sciences chemistry.chemical_compound 0302 clinical medicine Alu Elements Chromosome instability 030304 developmental biology Southern blot Palindromic sequence DNA Cruciform 0303 health sciences biology Inverted Repeat Sequences Chromosome Breakage biology.organism_classification Blotting Southern chemistry 030220 oncology & carcinogenesis Nucleic Acid Conformation Chromosomes Fungal Homologous recombination DNA |
| Zdroj: | Homologous Recombination ISBN: 9781071606438 Methods Mol Biol |
| DOI: | 10.1007/978-1-0716-0644-5_6 |
| Popis: | DNA repeats capable of adopting stable secondary structures are hotspots for double-strand break (DSB) formation and, hence, for homologous recombination and gross chromosomal rearrangements (GCR) in many prokaryotic and eukaryotic organisms, including humans. Here, we provide protocols for studying chromosomal instability triggered by hairpin- and cruciform-forming palindromic sequences in the budding yeast, Saccharomyces cerevisiae. First, we describe two sensitive genetic assays aimed to determine the recombinogenic potential of inverted repeats and their ability to induce GCRs. Then, we detail an approach to monitor chromosomal DSBs by Southern blot hybridization. Finally, we describe how to define the molecular structure of DSBs. We provide, as an example, the analysis of chromosomal fragility at a reporter system containing unstable Alu-inverted repeats. By using these approaches, any DNA sequence motif can be assessed for its breakage potential and ability to drive genome instability. |
| Databáze: | OpenAIRE |
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