title: Observation and Quantification of Telomere and Repetitive Sequences Using Fluorescence In Situ Hybridization (FISH) with PNA Probes in Caenorhabditis elegans.
abstract: Telomere is a ribonucleoprotein structure that protects chromosomal ends from aberrant fusion and degradation. Telomere length is maintained by telomerase or an alternative pathway, known as alternative lengthening of telomeres (ALT)(1). Recently, C. elegans has emerged as a multicellular model organism for the study of telomere and ALT(2). Visualization of repetitive sequences in the genome is critical in understanding the biology of telomeres. While telomere length can be measured by telomere restriction fragment assay or quantitative PCR, these methods only provide the averaged telomere length. On the contrary, fluorescence in situ hybridization (FISH) can provide the information of the individual telomeres in cells. Here, we provide protocols and representative results of the method to determine telomere length of C. elegans by fluorescent in situ hybridization. This method provides a simple, but powerful, in situ procedure that does not cause noticeable damage to morphology. By using fluorescently labeled peptide nucleic acid (PNA) and digoxigenin-dUTP-labeled probe, we were able to visualize two different repetitive sequences: telomere repeats and template of ALT (TALT) in C. elegans embryos and gonads.
authors: Institute of Molecular Biology and Genetics (IMBG), Seoul National University.
authors: Beomseok
authors: B
authors: Seo
authors: Institute of Molecular Biology and Genetics (IMBG), Seoul National University; Department of Biological Sciences, Seoul National University; Department of Biophysics and Chemical Biology, Seoul National University; elegans@snu.ac.kr.
authors: Junho
authors: J
authors: Lee
mesh_terms: D000818:Animals; D017173:Caenorhabditis elegans / Q000235:genetics* / Q000378:metabolism; D015342:DNA Probes / Q000737:chemistry / Q000235:genetics; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D020135:Peptide Nucleic Acids / Q000737:chemistry / Q000235:genetics; D012091:Repetitive Sequences, Nucleic Acid; D019098:Telomerase / Q000235:genetics / Q000378:metabolism; D016615:Telomere / Q000235:genetics / Q000378:metabolism*
chemical_list: D015342:DNA Probes; D020135:Peptide Nucleic Acids; D019098:Telomerase