title: A quick and simple FISH protocol with hybridization-sensitive fluorescent linear oligodeoxynucleotide probes.
abstract: Fluorescence in situ hybridization (FISH) is a powerful tool used in karyotyping, cytogenotyping, cancer diagnosis, species specification, and gene-expression analysis. Although widely used, conventional FISH protocols are cumbersome and time consuming. We have now developed a FISH method using exciton-controlled hybridization-sensitive fluorescent oligodeoxynucleotide (ECHO) probes. ECHO-FISH uses a 25-min protocol from fixation to mounting that includes no stringency washing steps. We use ECHO-FISH to detect both specific DNA and RNA sequences with multicolor probes. ECHO-FISH is highly reproducible, stringent, and compatible with other fluorescent cellular labeling techniques. The resolution allows detection of intranuclear speckles of poly(A) RNA in HeLa cells and dissociated hippocampal primary cultures, and mRNAs in the distal dendrites of hippocampal neurons. We also demonstrate detection of telomeric and centromeric DNA on metaphase mouse chromosomes. The simplicity of the ECHO-FISH method will likely accelerate cytogenetic and gene-expression analysis with high resolution.
authors.affiliation: Institute for Integrated Cell-Material Sciences, Kyoto University, Kyoto 606-8501, Japan. dwang@icems.kyoto-u.ac.jp
authors.forename: Dan Ohtan
authors.initials: DO
authors.lastname: Wang
authors.forename: Hitomi
authors.initials: H
authors.lastname: Matsuno
authors.forename: Shuji
authors.initials: S
authors.lastname: Ikeda
authors.forename: Akiko
authors.initials: A
authors.lastname: Nakamura
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Yanagisawa
authors.forename: Yasunori
authors.initials: Y
authors.lastname: Hayashi
authors.forename: Akimitsu
authors.initials: A
authors.lastname: Okamoto
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D004276:DNA, Satellite / Q000737:chemistry / Q000235:genetics; D053595:Embryonic Stem Cells / Q000737:chemistry / Q000378:metabolism; D005456:Fluorescent Dyes / Q000737:chemistry*; D020869:Gene Expression Profiling / Q000379:methods*; D006367:HeLa Cells; D006624:Hippocampus / Q000737:chemistry; D006801:Humans; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D051379:Mice; D009838:Oligodeoxyribonucleotides / Q000737:chemistry*; D012333:RNA, Messenger / Q000032:analysis*; D016615:Telomere / Q000737:chemistry / Q000235:genetics
chemical_list: D004276:DNA, Satellite; D005456:Fluorescent Dyes; D009838:Oligodeoxyribonucleotides; D012333:RNA, Messenger
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