A fast, low cost, and highly efficient fluorescent DNA labeling method using methyl green
Resumen:
The increasing need for multiple-labeling of cells and whole organisms for fluorescence microscopy has led to the development of hundreds of fluorophores that either directly recognize target molecules or organelles, or are attached to antibodies or other molecular probes. DNA labeling is essential to study nuclear-chromosomal structure, as well as for gel staining, but also as a usual counterstain in immunofluorescence, FISH or cytometry. However, there are currently few reliable red to far-red-emitting DNA stains that can be used. We describe herein an extremely simple, inexpensive and robust method for DNA labeling of cells and electrophoretic gels using the very well-known histological stain methyl green (MG). MG used in very low concentrations at physiological pH proved to have relatively narrow excitation and emission spectra, with peaks at 633 and 677 nm, respectively, and a very high resistance to photobleaching. It can be used in combination with other common DNA stains or antibodies without any visible interference or bleed-through. In electrophoretic gels, MG also labeled DNA in a similar way to ethidium bromide, but, as expected, it did not label RNA. Moreover, we show here that MG fluorescence can be used as a stain for direct measuring of viability by both microscopy and flow cytometry, with full correlation to ethidium bromide staining. MG is thus a very convenient alternative to currently used red-emitting DNA stains.
2014 | |
Methyl green DNA Flow cytometry Confocal microscopy Electrophoresis Fluorescence |
|
Inglés | |
Universidad de la República | |
COLIBRI | |
https://hdl.handle.net/20.500.12008/22055 | |
Acceso abierto | |
Licencia Creative Commons Atribución – No Comercial – Sin Derivadas (CC –BY-NC-ND 4.0) |
_version_ | 1807522780187459584 |
---|---|
author | Prieto Mena, Daniel |
author2 | Aparicio Díaz, Hector Gonzalo Morande, Pablo E. Zolessi, Flavio R. |
author2_role | author author author |
author_facet | Prieto Mena, Daniel Aparicio Díaz, Hector Gonzalo Morande, Pablo E. Zolessi, Flavio R. |
author_role | author |
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collection | COLIBRI |
dc.contributor.filiacion.es.fl_str_mv | Prieto Mena, Daniel. IIBCE Aparicio Díaz, Hector Gonzalo. Universidad de la República (Uruguay). Facultad de Ciencias. Instituto de Biología Morande, Pablo E. Instituto Pasteur (Montevideo) Zolessi, Flavio R. Universidad de la República (Uruguay). Facultad de Ciencias. Instituto de Biología |
dc.creator.none.fl_str_mv | Prieto Mena, Daniel Aparicio Díaz, Hector Gonzalo Morande, Pablo E. Zolessi, Flavio R. |
dc.date.accessioned.none.fl_str_mv | 2019-10-02T22:12:05Z |
dc.date.available.none.fl_str_mv | 2019-10-02T22:12:05Z |
dc.date.issued.es.fl_str_mv | 2014 |
dc.date.submitted.es.fl_str_mv | 20190930 |
dc.description.abstract.none.fl_txt_mv | The increasing need for multiple-labeling of cells and whole organisms for fluorescence microscopy has led to the development of hundreds of fluorophores that either directly recognize target molecules or organelles, or are attached to antibodies or other molecular probes. DNA labeling is essential to study nuclear-chromosomal structure, as well as for gel staining, but also as a usual counterstain in immunofluorescence, FISH or cytometry. However, there are currently few reliable red to far-red-emitting DNA stains that can be used. We describe herein an extremely simple, inexpensive and robust method for DNA labeling of cells and electrophoretic gels using the very well-known histological stain methyl green (MG). MG used in very low concentrations at physiological pH proved to have relatively narrow excitation and emission spectra, with peaks at 633 and 677 nm, respectively, and a very high resistance to photobleaching. It can be used in combination with other common DNA stains or antibodies without any visible interference or bleed-through. In electrophoretic gels, MG also labeled DNA in a similar way to ethidium bromide, but, as expected, it did not label RNA. Moreover, we show here that MG fluorescence can be used as a stain for direct measuring of viability by both microscopy and flow cytometry, with full correlation to ethidium bromide staining. MG is thus a very convenient alternative to currently used red-emitting DNA stains. |
dc.description.es.fl_txt_mv | Preprint presentado a Springer |
dc.format.mimetype.es.fl_str_mv | application/pdf |
dc.identifier.citation.es.fl_str_mv | Prieto, D., y otros. "A fast, low cost, and highly efficient fluorescent DNA labeling method using methyl green" [Preprint]. Publicado en: Histochemistry and Cell Biology, 2014, 142 (3): 335-345. doi: 10.1007/s00418-014-1215-0 |
dc.identifier.uri.none.fl_str_mv | https://hdl.handle.net/20.500.12008/22055 |
dc.language.iso.none.fl_str_mv | en eng |
dc.rights.license.none.fl_str_mv | Licencia Creative Commons Atribución – No Comercial – Sin Derivadas (CC –BY-NC-ND 4.0) |
dc.rights.none.fl_str_mv | info:eu-repo/semantics/openAccess |
dc.source.none.fl_str_mv | reponame:COLIBRI instname:Universidad de la República instacron:Universidad de la República |
dc.subject.es.fl_str_mv | Methyl green DNA Flow cytometry Confocal microscopy Electrophoresis Fluorescence |
dc.title.none.fl_str_mv | A fast, low cost, and highly efficient fluorescent DNA labeling method using methyl green |
dc.type.en.fl_str_mv | Preprint |
dc.type.none.fl_str_mv | info:eu-repo/semantics/preprint |
dc.type.version.none.fl_str_mv | info:eu-repo/semantics/submittedVersion |
description | Preprint presentado a Springer |
eu_rights_str_mv | openAccess |
format | preprint |
id | COLIBRI_d415ee6a1333a2a6e2d1dad731079f2b |
identifier_str_mv | Prieto, D., y otros. "A fast, low cost, and highly efficient fluorescent DNA labeling method using methyl green" [Preprint]. Publicado en: Histochemistry and Cell Biology, 2014, 142 (3): 335-345. doi: 10.1007/s00418-014-1215-0 |
instacron_str | Universidad de la República |
institution | Universidad de la República |
instname_str | Universidad de la República |
language | eng |
language_invalid_str_mv | en |
network_acronym_str | COLIBRI |
network_name_str | COLIBRI |
oai_identifier_str | oai:colibri.udelar.edu.uy:20.500.12008/22055 |
publishDate | 2014 |
reponame_str | COLIBRI |
repository.mail.fl_str_mv | mabel.seroubian@seciu.edu.uy |
repository.name.fl_str_mv | COLIBRI - Universidad de la República |
repository_id_str | 4771 |
rights_invalid_str_mv | Licencia Creative Commons Atribución – No Comercial – Sin Derivadas (CC –BY-NC-ND 4.0) |
spelling | Prieto Mena, Daniel. IIBCEAparicio Díaz, Hector Gonzalo. Universidad de la República (Uruguay). Facultad de Ciencias. Instituto de BiologíaMorande, Pablo E. Instituto Pasteur (Montevideo)Zolessi, Flavio R. Universidad de la República (Uruguay). Facultad de Ciencias. Instituto de Biología2019-10-02T22:12:05Z2019-10-02T22:12:05Z201420190930Prieto, D., y otros. "A fast, low cost, and highly efficient fluorescent DNA labeling method using methyl green" [Preprint]. Publicado en: Histochemistry and Cell Biology, 2014, 142 (3): 335-345. doi: 10.1007/s00418-014-1215-0https://hdl.handle.net/20.500.12008/22055Preprint presentado a SpringerThe increasing need for multiple-labeling of cells and whole organisms for fluorescence microscopy has led to the development of hundreds of fluorophores that either directly recognize target molecules or organelles, or are attached to antibodies or other molecular probes. DNA labeling is essential to study nuclear-chromosomal structure, as well as for gel staining, but also as a usual counterstain in immunofluorescence, FISH or cytometry. However, there are currently few reliable red to far-red-emitting DNA stains that can be used. We describe herein an extremely simple, inexpensive and robust method for DNA labeling of cells and electrophoretic gels using the very well-known histological stain methyl green (MG). MG used in very low concentrations at physiological pH proved to have relatively narrow excitation and emission spectra, with peaks at 633 and 677 nm, respectively, and a very high resistance to photobleaching. It can be used in combination with other common DNA stains or antibodies without any visible interference or bleed-through. In electrophoretic gels, MG also labeled DNA in a similar way to ethidium bromide, but, as expected, it did not label RNA. Moreover, we show here that MG fluorescence can be used as a stain for direct measuring of viability by both microscopy and flow cytometry, with full correlation to ethidium bromide staining. MG is thus a very convenient alternative to currently used red-emitting DNA stains.Made available in DSpace on 2019-10-02T22:12:05Z (GMT). No. of bitstreams: 5 101007s0041801412150.pdf: 3672120 bytes, checksum: 6b58577ddb18cbbba34972c6cd796920 (MD5) license_text: 38300 bytes, checksum: 098d76773c7b7afafb04cabc04ea8a56 (MD5) license_url: 47 bytes, checksum: 966d4a1cc97b2c4389b5142dd97d3c7f (MD5) license_rdf: 9754 bytes, checksum: ffcba5f515f45166c8d3bb6aa02e3123 (MD5) license.txt: 4194 bytes, checksum: 7f2e2c17ef6585de66da58d1bfa8b5e1 (MD5) Previous issue date: 2014application/pdfenengLas obras depositadas en el Repositorio se rigen por la Ordenanza de los Derechos de la Propiedad Intelectual de la Universidad De La República. (Res. Nº 91 de C.D.C. de 8/III/1994 – D.O. 7/IV/1994) y por la Ordenanza del Repositorio Abierto de la Universidad de la República (Res. Nº 16 de C.D.C. de 07/10/2014)info:eu-repo/semantics/openAccessLicencia Creative Commons Atribución – No Comercial – Sin Derivadas (CC –BY-NC-ND 4.0)Methyl greenDNAFlow cytometryConfocal microscopyElectrophoresisFluorescenceA fast, low cost, and highly efficient fluorescent DNA labeling method using methyl greenPreprintinfo:eu-repo/semantics/preprintinfo:eu-repo/semantics/submittedVersionreponame:COLIBRIinstname:Universidad de la Repúblicainstacron:Universidad de la RepúblicaPrieto Mena, DanielAparicio Díaz, Hector GonzaloMorande, Pablo E.Zolessi, Flavio 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://udelar.edu.uy/https://www.colibri.udelar.edu.uy/oai/requestmabel.seroubian@seciu.edu.uyUruguayopendoar:47712024-07-25T14:28:09.730870COLIBRI - Universidad de la Repúblicafalse |
spellingShingle | A fast, low cost, and highly efficient fluorescent DNA labeling method using methyl green Prieto Mena, Daniel Methyl green DNA Flow cytometry Confocal microscopy Electrophoresis Fluorescence |
status_str | submittedVersion |
title | A fast, low cost, and highly efficient fluorescent DNA labeling method using methyl green |
title_full | A fast, low cost, and highly efficient fluorescent DNA labeling method using methyl green |
title_fullStr | A fast, low cost, and highly efficient fluorescent DNA labeling method using methyl green |
title_full_unstemmed | A fast, low cost, and highly efficient fluorescent DNA labeling method using methyl green |
title_short | A fast, low cost, and highly efficient fluorescent DNA labeling method using methyl green |
title_sort | A fast, low cost, and highly efficient fluorescent DNA labeling method using methyl green |
topic | Methyl green DNA Flow cytometry Confocal microscopy Electrophoresis Fluorescence |
url | https://hdl.handle.net/20.500.12008/22055 |