Note to users. If you're seeing this message, it means that your browser cannot find this page's style/presentation instructions -- or possibly that you are using a browser that does not support current Web standards. Find out more about why this message is appearing, and what you can do to make your experience of our site the best it can be.

Site Tools

  • AAAS
  • Subscribe
  • Feedback

Site Search

Search Advanced

Science 24 April 1998:
Vol. 280. no. 5363, pp. 585 - 590
DOI: 10.1126/science.280.5363.585
Prev | Table of Contents | Next

Reports

Visualization of Single RNA Transcripts in Situ

Andrea M. Femino, Fredric S. Fay, dagger Kevin Fogarty, Robert H. Singer *

Fluorescence in situ hybridization (FISH) and digital imaging microscopy were modified to allow detection of single RNA molecules. Oligodeoxynucleotide probes were synthesized with five fluorochromes per molecule, and the light emitted by a single probe was calibrated. Points of light in exhaustively deconvolved images of hybridized cells gave fluorescent intensities and distances between probes consistent with single messenger RNA molecules. Analysis of beta -actin transcription sites after serum induction revealed synchronous and cyclical transcription from single genes. The rates of transcription initiation and termination and messenger RNA processing could be determined by positioning probes along the transcription unit. This approach extends the power of FISH to yield quantitative molecular information on a single cell.

A. M. Femino and R. H. Singer, Department of Anatomy and Structural Biology and Cell Biology, Albert Einstein College of Medicine, Bronx, NY 10461, USA.
F. S. Fay and K. Fogarty, Biomedical Imaging Facility and Department of Physiology, University of Massachusetts Medical School, Worcester, MA 01655, USA.
*   To whom correspondence should be addressed.

dagger    Deceased.

Read the Full Text


THIS ARTICLE HAS BEEN CITED BY OTHER ARTICLES:
Translation inhibition reveals interaction of 2'-deoxy and 2'-O-methyl molecular beacons with mRNA targets in living cells.
N. Nitin, W. J. Rhee, and G. Bao (2009)
Nucleic Acids Res.
   Abstract »    Full Text »    PDF »
Imaging individual microRNAs in single mammalian cells in situ.
J. Lu and A. Tsourkas (2009)
Nucleic Acids Res.
   Abstract »    Full Text »    PDF »
The Dynamics of Mammalian P Body Transport, Assembly, and Disassembly In Vivo.
A. Aizer, Y. Brody, L. W. Ler, N. Sonenberg, R. H. Singer, and Y. Shav-Tal (2008)
Mol. Biol. Cell 19, 4154-4166
   Abstract »    Full Text »    PDF »
Single-Cell Transcription Site Activation Predicts Chemotherapy Response in Human Colorectal Tumors.
R. C. Pezo, S. J. Gandhi, L. A. Shirley, R. G. Pestell, L. H. Augenlicht, and R. H. Singer (2008)
Cancer Res. 68, 4977-4982
   Abstract »    Full Text »    PDF »
RNA polymerase II activity is located on the surface of protein-rich transcription factories.
C. H. Eskiw, A. Rapp, D. R. F. Carter, and P. R. Cook (2008)
J. Cell Sci. 121, 1999-2007
   Abstract »    Full Text »    PDF »
Single-molecule biophysics: at the interface of biology, physics and chemistry.
A. A Deniz, S. Mukhopadhyay, and E. A Lemke (2008)
J R Soc Interface 5, 15-45
   Abstract »    Full Text »    PDF »
Imaging Real-Time Gene Expression in Living Systems with Single-Transcript Resolution: Image Analysis of Single mRNA Transcripts.
A. L. Wells, J. S. Condeelis, R. H. Singer, and D. Zenklusen (2007)
CSH Protocols 2007, pdb.prot4871
   Abstract »    Full Text »
The transcriptional cycle of HIV-1 in real-time and live cells.
S. Boireau, P. Maiuri, E. Basyuk, M. de la Mata, A. Knezevich, B. Pradet-Balade, V. Backer, A. Kornblihtt, A. Marcello, and E. Bertrand (2007)
J. Cell Biol. 179, 291-304
   Abstract »    Full Text »    PDF »
Bsr, a Nuclear-retained RNA with Monoallelic Expression.
H. Royo, E. Basyuk, V. Marty, M. Marques, E. Bertrand, and J. Cavaille (2007)
Mol. Biol. Cell 18, 2817-2827
   Abstract »    Full Text »    PDF »
Single-molecule Chemistry and Biology Special Feature: New directions in single-molecule imaging and analysis.
W. E. Moerner (2007)
PNAS 104, 12596-12602
   Abstract »    Full Text »    PDF »
Noise in Gene Expression Determines Cell Fate in Bacillus subtilis.
H. Maamar, A. Raj, and D. Dubnau (2007)
Science 317, 526-529
   Abstract »    Full Text »    PDF »
Mitochondria-associated Yeast mRNAs and the Biogenesis of Molecular Complexes.
M. Garcia, X. Darzacq, T. Delaveau, L. Jourdren, R. H. Singer, and C. Jacq (2007)
Mol. Biol. Cell 18, 362-368
   Abstract »    Full Text »    PDF »
Visualization of mRNA translation in living cells.
A. J. Rodriguez, S. M. Shenoy, R. H. Singer, and J. Condeelis (2006)
J. Cell Biol. 175, 67-76
   Abstract »    Full Text »    PDF »
Many expressed genes in bacteria and yeast are transcribed only once per cell cycle.
M. Bon, S. J. McGowan, and P. R. Cook (2006)
FASEB J 20, 1721-1723
   Abstract »    Full Text »    PDF »
Assembling an intermediate filament network by dynamic cotranslation.
L. Chang, Y. Shav-Tal, T. Trcek, R. H. Singer, and R. D. Goldman (2006)
J. Cell Biol. 172, 747-758
   Abstract »    Full Text »    PDF »
Cell Cycle-dependent Recruitment of Telomerase RNA and Cajal Bodies to Human Telomeres.
B. E. Jady, P. Richard, E. Bertrand, and T. Kiss (2006)
Mol. Biol. Cell 17, 944-954
   Abstract »    Full Text »    PDF »
Stochastic simulation of the mammalian circadian clock.
D. B. Forger and C. S. Peskin (2005)
PNAS 102, 321-324
   Abstract »    Full Text »    PDF »
Dynamics of Single mRNPs in Nuclei of Living Cells.
Y. Shav-Tal, X. Darzacq, S. M. Shenoy, D. Fusco, S. M. Janicki, D. L. Spector, and R. H. Singer (2004)
Science 304, 1797-1800
   Abstract »    Full Text »    PDF »
Dual FRET molecular beacons for mRNA detection in living cells.
P. J. Santangelo, B. Nix, A. Tsourkas, and G. Bao (2004)
Nucleic Acids Res. 32, e57
   Abstract »    Full Text »    PDF »
Peptide-linked molecular beacons for efficient delivery and rapid mRNA detection in living cells.
N. Nitin, P. J. Santangelo, G. Kim, S. Nie, and G. Bao (2004)
Nucleic Acids Res. 32, e58
   Abstract »    Full Text »    PDF »
Induction of CYP1A1 in Primary Rat Hepatocytes by 3,3',4,4',5-Pentachlorobiphenyl: Evidence for a Switch Circuit Element.
C. T. French, W. H. Hanneman, L. S. Chubb, R. E. Billings, and M. E. Andersen (2004)
Toxicol. Sci. 78, 276-286
   Abstract »    Full Text »    PDF »
Widespread cytoplasmic mRNA transport in yeast: Identification of 22 bud-localized transcripts using DNA microarray analysis.
K. A. Shepard, A. P. Gerber, A. Jambhekar, P. A. Takizawa, P. O. Brown, D. Herschlag, J. L. DeRisi, and R. D. Vale (2003)
PNAS 100, 11429-11434
   Abstract »    Full Text »    PDF »
Fluorescence in situ hybridization: past, present and future.
J. M. Levsky and R. H. Singer (2003)
J. Cell Sci. 116, 2833-2838
   Abstract »    Full Text »    PDF »
Application of Gene Expression Profiling to Colon Cell Maturation, Transformation and Chemoprevention.
L. H. Augenlicht, A. Velcich, L. Klampfer, J. Huang, G. Corner, M. Aranes, C. Laboisse, B. Rigas, M. Lipkin, K. Yang, et al. (2003)
J. Nutr. 133, 2410S-2416
   Abstract »    Full Text »    PDF »
Cloning, Localization, and Axonemal Function of Tetrahymena Centrin.
C. Guerra, Y. Wada, V. Leick, A. Bell, and P. Satir (2003)
Mol. Biol. Cell 14, 251-261
   Abstract »    Full Text »
The transcription cycle of RNA polymerase II in living cells.
H. Kimura, K. Sugaya, and P. R. Cook (2002)
J. Cell Biol. 159, 777-782
   Abstract »    Full Text »    PDF »
Experience-Dependent Coincident Expression of the Effector Immediate-Early Genes Arc and Homer 1a in Hippocampal and Neocortical Neuronal Networks.
A. Vazdarjanova, B. L. McNaughton, C. A. Barnes, P. F. Worley, and J. F. Guzowski (2002)
J. Neurosci. 22, 10067-10071
   Abstract »    Full Text »    PDF »
Novel Detection and Differential Utilization of a c-myc Transcriptional Block in Colon Cancer Chemoprevention.
A. J. Wilson, A. Velcich, D. Arango, A. R. Kurland, S. M. Shenoy, R. C. Pezo, J. M. Levsky, R. H. Singer, and L. H. Augenlicht (2002)
Cancer Res. 62, 6006-6010
   Abstract »    Full Text »    PDF »
Detection of Viral Infection and Gene Expression in Clinical Tissue Specimens Using Branched DNA (bDNA) In Situ Hybridization.
D. Kenny, L.-P. Shen, and J. A. Kolberg (2002)
J. Histochem. Cytochem. 50, 1219-1227
   Abstract »    Full Text »    PDF »
Allele-specific expression analysis by RNA-FISH demonstrates preferential maternal expression of UBE3A and imprint maintenance within 15q11- q13 duplications.
L. B.K. Herzing, E. H. Cook Jr, and D. H. Ledbetter (2002)
Hum. Mol. Genet. 11, 1707-1718
   Abstract »    Full Text »    PDF »
Kinetics of HCMV immediate early mRNA expression in stably transfected fibroblasts.
S. P. Snaar, P. Verdijk, H. J. Tanke, and R. W. Dirks (2002)
J. Cell Sci. 115, 321-328
   Abstract »    Full Text »    PDF »
Efficient Hepatitis Delta Virus RNA Replication in Avian Cells Requires a Permissive Factor(s) from Mammalian Cells.
Y.-T. Liu, R. Brazas, and D. Ganem (2001)
J. Virol. 75, 7489-7493
   Abstract »    Full Text »    PDF »
The physiological significance of beta -actin mRNA localization in determining cell polarity and directional motility.
E. A. Shestakova, R. H. Singer, and J. Condeelis (2001)
PNAS 98, 7045-7050
   Abstract »    Full Text »    PDF »
Optimizing the detection of nascent transcripts by RNA fluorescence in situ hybridization.
C. D. van Raamsdonk and S. M. Tilghman (2001)
Nucleic Acids Res. 29, e42
   Abstract »    Full Text »    PDF »
Computational adaptive optics for live three-dimensional biological imaging.
Z. Kam, B. Hanser, M. G. L. Gustafsson, D. A. Agard, and J. W. Sedat (2001)
PNAS 98, 3790-3795
   Abstract »    Full Text »    PDF »
Probability in transcriptional regulation and its implications for leukocyte differentiation and inducible gene expression.
D. A. Hume (2000)
Blood 96, 2323-2328
   Abstract »    Full Text »    PDF »
The balance sheet for transcription: an analysis of nuclear RNA metabolism in mammalian cells.
D. A. JACKSON, A. POMBO, and F. IBORRA (2000)
FASEB J 14, 242-254
   Abstract »    Full Text »
Large-scale chromatin organization of the major histocompatibility complex and other regions of human chromosome 6 and its response to interferon in interphase nuclei.
E. Volpi, E Chevret, T Jones, R Vatcheva, J Williamson, S Beck, R. Campbell, M Goldsworthy, S. Powis, J Ragoussis, et al. (2000)
J. Cell Sci. 113, 1565-1576
   Abstract »    PDF »
Effects of macromolecular transport and stochastic fluctuations on dynamics of genetic regulatory systems.
P. Smolen, D. A. Baxter, and J. H. Byrne (1999)
Am J Physiol Cell Physiol 277, C777-C790
   Abstract »    Full Text »    PDF »
The Organization of Replication and Transcription.
P. R. Cook (1999)
Science 284, 1790-1795
   Abstract »    Full Text »
Single-molecule Enzymology.
X. S. Xie and H. P. Lu (1999)
J. Biol. Chem. 274, 15967-15970
   Full Text »    PDF »
Amplification Methods to Increase the Sensitivity of In Situ Hybridization: Play CARD(S).
E. J.M. Speel, A. H.N. Hopman, and P. Komminoth (1999)
J. Histochem. Cytochem. 47, 281-288
   Abstract »    Full Text »
The travels of mRNAs through all cells large and small.
G. J. BASSELL, Y. OLEYNIKOV, and R. H. SINGER (1999)
FASEB J 13, 447-454
   Full Text »
3-D organization of ribosomal transcription units after DRB inhibition of RNA polymerase II transcription.
S. Panse, C Masson, L Heliot, J. Chassery, H. Junera, and D Hernandez-Verdun (1999)
J. Cell Sci. 112, 2145-2154
   Abstract »    PDF »
Ultrastructural Localization of ß-Actin and Amphoterin mRNA in Cultured Cells: Application of Tyramide Signal Amplification and Comparison of Detection Methods.
E.-L. Punnonen, C. Fages, J. Wartiovaara, and H. Rauvala (1999)
J. Histochem. Cytochem. 47, 99-112
   Abstract »    Full Text »
Structure and Function in the Nucleus.
A. I. Lamond and W. C. Earnshaw (1998)
Science 280, 547-553
   Abstract »    Full Text »


To Advertise     Find Products

Science. ISSN 0036-8075 (print), 1095-9203 (online)