Abstract
Oligonucleotide probes labeled with fluorescent dyes are used in a variety of in situ applications to detect specific DNA or RNA molecules. It has been described that probe fluorescence might be quenched upon hybridization in a sequence specific way. Here, a set of 17 oligonuleotides labeled with 6-carboxyfluorescein was used to examine the relevance of nucleotide specific quenching for fluorescence in situ hybridization (FISH) to whole fixed bacterial cells. Probes quenched upon hybridization to a guanine-rich region of purified RNA in solution were not quenched upon FISH. Among other factors the high protein concentration within cells may prevent quenching of probe fluorescence in situ.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 565-572 |
| Number of pages | 8 |
| Journal | Systematic and Applied Microbiology |
| Volume | 27 |
| Issue number | 5 |
| DOIs | |
| State | Published - Sep 2004 |
| Externally published | Yes |
Bibliographical note
Funding Information:We thank Markus Sauer (Institute of Physical Chemistry, University of Heidelberg) for helpful discussions on the physics of quenching, and Werner Wosniok (University of Bremen) for advice on statistics. This work was supported by grants from the Deutsche Forschungsgemeinschaft (Am73/3-1), the Fonds der chemischen Industrie, and the Max Planck Society.
Copyright:
Copyright 2017 Elsevier B.V., All rights reserved.
Keywords
- 6-carboxyfluorescein (6-FAM)
- Fluorescence in situ hybridization (FISH)
- Fluorescent dyes
- Nucleobase-specific quenching
- Oligonucleotide probes
- Photoinduced electron transfer