DOI: 10.7554/elife.01775. Single-molecule tracking of the transcription cycle by sub-second RNA detection

Single-molecule tracking of the transcription cycle by sub-second RNA detection

10.7554/elife.01775

Crossref journal-article

eLife Sciences Publications, Ltd

eLife (4374)

Abstract

Transcription is an inherently stochastic, noisy, and multi-step process, in which fluctuations at every step can cause variations in RNA synthesis, and affect physiology and differentiation decisions in otherwise identical cells. However, it has been an experimental challenge to directly link the stochastic events at the promoter to transcript production. Here we established a fast fluorescence in situ hybridization (fastFISH) method that takes advantage of intrinsically unstructured nucleic acid sequences to achieve exceptionally fast rates of specific hybridization (∼10e7 M−1s−1), and allows deterministic detection of single nascent transcripts. Using a prototypical RNA polymerase, we demonstrated the use of fastFISH to measure the kinetic rates of promoter escape, elongation, and termination in one assay at the single-molecule level, at sub-second temporal resolution. The principles of fastFISH design can be used to study stochasticity in gene regulation, to select targets for gene silencing, and to design nucleic acid nanostructures.

Bibliography

Zhang, Z., Revyakin, A., Grimm, J. B., Lavis, L. D., & Tjian, R. (2014). Single-molecule tracking of the transcription cycle by sub-second RNA detection. ELife, 3. CLOCKSS.

Authors 5

Zhengjian Zhang (first)
Andrey Revyakin (additional)
Jonathan B Grimm (additional)
Luke D Lavis (additional)
Robert Tjian (additional)

References 67 Referenced 70

10.1016/0022-2836(88)90457-3 / Journal of Molecular Biology / Transcription termination in Escherichia coli. Measurement of the rate of enzyme release from Rho-independent terminators by Arndt (1988)
10.1146/annurev.biophys.35.010406.150153 / Annual Review of Biophysics and Biomolecular Structure / Single-molecule analysis of RNA polymerase transcription by Bai (2006)
10.1073/pnas.0403884101 / Proceedings of the National Academy of Sciences of the United States of America / tRNA dynamics on the ribosome during translation by Blanchard (2004)
10.1016/S0021-9258(17)31506-5 / Journal of Biological Chemistry / Characterization of a set of T7 RNA polymerase active site mutants by Bonner (1994)
10.1126/science.1218716 / Science / Opening and closing of the bacterial RNA polymerase clamp by Chakraborty (2012)
10.1016/S0006-3495(95)80095-0 / Biophysical Journal / The biophysics of DNA hybridization with immobilized oligonucleotide probes by Chan (1995)
10.1016/j.cell.2012.06.006 / Cell / A movie of RNA polymerase ii transcription by Cheung (2012)
10.1128/JB.180.12.3019-3025.1998 / Journal of Bacteriology / RNA polymerase-promoter interactions: the comings and goings of RNA Polymerase by DeHaseth (1998)
10.1103/PhysRevLett.89.248102 / Physical Review Letters / Stretching single stranded DNA, a model polyelectrolyte by Dessinges (2002)
10.1126/science.1083219 / Science / Cooperation between RNA polymerase molecules in transcription elongation by Epshtein (2003)
10.1529/biophysj.106.084004 / Biophysical Journal / Viewing dynamic assembly of molecular complexes by multi-wavelength single-molecule fluorescence by Friedman (2006)
10.1016/j.cell.2012.01.018 / Cell / Mechanism of transcription initiation at an activator-dependent promoter defined by single-molecule observation by Friedman (2012)
10.1073/pnas.1300221110 / Proceedings of the National Academy of Sciences of the United States of America / RNA polymerase approaches its promoter without long-range sliding along DNA by Friedman (2013)
10.1038/nature05701 / Nature / Backtracking determines the force sensitivity of RNAP II in a factor-dependent manner by Galburt (2007)
10.1093/nar/gkl422 / Nucleic Acids Research / Secondary structure effects on DNA hybridization kinetics: a solution versus surface comparison by Gao (2006)
10.1016/S0021-9258(19)42709-9 / Journal of Biological Chemistry / Characterization of T7-specific ribonucleic acid polymerase: IV. Resolution of the major in vitro transcripts by gel electrophoresis by Golomb (1974)
10.1016/S0006-3495(02)75672-5 / Biophysical Journal / Magnetic tweezers: micromanipulation and force measurement at the molecular level by Gosse (2002)
10.1093/nar/gkn188 / Nucleic Acids Research / The Vienna RNA websuite by Gruber (2008)
10.1016/j.cell.2011.07.026 / Cell / Stochastic state transitions give rise to phenotypic equilibrium in populations of cancer cells by Gupta (2011)
10.1016/j.molcel.2009.09.005 / Molecular Cell / Transcription dynamics by Hager (2009)
10.1146/annurev.biochem.77.061906.092014 / Annual Review of Biochemistry / Super resolution fluorescence microscopy by Huang (2010)
10.1006/jmbi.2000.4150 / Journal of Molecular Biology / T7 RNA polymerase elongation complex structure and movement by Huang (2000)
10.1074/jbc.270.5.2290 / Journal of Biological Chemistry / RNA polymerase II ternary complexes may become arrested after transcribing to within 10 bases of the end of linear templates by Izban (1995)
10.1021/ja025678o / Journal of the American Chemical Society / Intramolecular dimers: a new strategy to fluorescence quenching in dual-labeled oligonucleotide probes by Johansson (2002)
10.1126/science.1131399 / Science / Initial transcription by RNA polymerase proceeds through a DNA-scrunching mechanism by Kapanidis (2006)
10.1103/PhysRevA.36.842 / Physical Review A / Easily calculable measure for the complexity of spatiotemporal patterns by Kaspar (1987)
10.1021/ja0041324 / Journal of the American Chemical Society / Hybridization of DNA and PNA molecular beacons to single-stranded and double-stranded DNA targets by Kuhn (2002)
10.1021/ja016310e / Journal of the American Chemical Society / Effect of secondary structure on the thermodynamics and kinetics of PNA hybridization to DNA hairpins by Kushon (2001)
10.1021/bi00163a013 / Biochemistry / Implication of RNA structure on antisense oligonucleotide hybridization kinetics by Lima (1992)
10.1038/embor.2012.31 / EMBO Reports / Transcription goes digital by Lionnet (2012)
10.1126/science.1235441 / Science / Transcription under torsion by Ma (2013)
10.1093/nar/gnf121 / Nucleic Acids Research / Efficiencies of fluorescence resonance energy transfer and contact-mediated quenching in oligonucleotide probes by Marras (2002)
10.1021/bi00384a006 / Biochemistry / Kinetic analysis of T7 RNA polymerase-promoter interactions with small synthetic promoters by Martin (1987)
10.1021/bi00067a017 / Biochemistry / Kinetic analysis of T7 RNA polymerase transcription initiation from promoters containing single-stranded regions by Maslak (1993)
10.1093/nar/15.21.8783 / Nucleic Acids Research / Oligoribonucleotide synthesis using T7 RNA polymerase and synthetic DNA templates by Milligan (1987)
10.1021/bc00020a001 / Bioconjugate Chemistry / Cyanine dye labeling reagents: sulfoindocyanine succinimidyl esters by Mujumdar (1993)
10.1016/S0959-440X(02)00005-2 / Current Opinion in Structural Biology / Bacterial RNA polymerases: the wholo story by Murakami (2003)
10.1016/S0092-8674(03)00845-6 / Cell / Ubiquitous transcriptional pausing is independent of RNA polymerase backtracking by Neuman (2003)
10.1182/blood.V81.11.2844.2844 / Blood / Differentiation and proliferation of hematopoietic stem cells by Ogawa (1993)
10.1093/nar/gkh466 / Nucleic Acids Research / Complexity: an internet resource for analysis of DNA sequence complexity by Orlov (2004)
10.1002/9780470559277.ch110148 / Current Protocols in Chemical Biology / Copper-catalyzed azide-alkyne click chemistry for bioconjugation by Presolski (2011)
10.1146/annurev.biophys.37.032807.125928 / Annual Review of Biophysics / Single-molecule approaches to stochastic gene expression by Raj (2009)
10.1073/pnas.90.8.3147 / Proceedings of the National Academy of Sciences of the United States of America / T7 RNA polymerase mutants with altered promoter specificities by Raskin (1993)
10.1126/science.1131398 / Science / Abortive initiation and productive initiation by RNA polymerase involve DNA scrunching by Revyakin (2006)
10.1101/gad.194936.112 / Genes & Development / Transcription initiation by human RNA polymerase II visualized at single-molecule resolution by Revyakin (2012)
10.1016/j.jmb.2012.12.015 / Journal of Molecular Biology / The transcription bubble of the RNA polymerase-promoter open complex exhibits conformational heterogeneity and millisecond-scale dynamics: implications for transcription start-site selection by Robb (2013)
10.1038/nature04586 / Nature / Folding DNA to create nanoscale shapes and patterns by Rothemund (2006)
10.1002/9780470316849 / Multivariate density estimation: theory, practice, and visualization (Wiley series in probability and statistics) by Scott (1992)
10.1038/nature02191 / Nature / Backtracking by single RNA polymerase molecules observed at near-base-pair resolution by Shaevitz (2003)
10.1074/jbc.M310471200 / Journal of Biological Chemistry / Promoter binding, initiation, and elongation by bacteriophage T7 RNA polymerase. A single-molecule view of the transcription cycle by Skinner (2004)
10.1126/science.271.5250.795 / Science / Overstretching B-DNA: the elastic response of individual double-stranded and single-stranded DNA molecules by Smith (1996)
10.1074/jbc.M201600200 / Journal of Biological Chemistry / The +2 NTP binding drives open complex formation in T7 RNA polymerase by Stano (2002)
10.1021/bi052292s / Biochemistry / Rapid binding of T7 RNA polymerase is followed by simultaneous bending and opening of the promoter DNA by Tang (2006)
10.1073/pnas.0906979106 / Proceedings of the National Academy of Sciences of the United States of America / Real-time observation of the transition from transcription initiation to elongation of the RNA polymerase by Tang (2009)
10.1529/biophysj.107.125096 / Biophysical Journal / T7 RNA polymerase studied by force measurements varying cofactor concentration by Thomen (2008)
10.1093/nar/gkg212 / Nucleic Acids Research / Hybridization kinetics and thermodynamics of molecular beacons by Tsourkas (2003)
10.1038/nsmb.2472 / Nature Structure Molecular Biology / The promoter-search mechanism of Escherichia coli RNA polymerase is dominated by three-dimensional diffusion by Wang (2012)
10.1089/oli.2004.14.167 / Oligonucleotides / Computational identification of antisense oligonucleotides that rapidly hybridize to RNA by Wang (2004)
10.1126/science.282.5390.902 / Science / Force and velocity measured for single molecules of RNA polymerase by Wang (1998)
10.1126/science.283.5408.1676 / Science / Fluorescence spectroscopy of single biomolecules by Weiss (1999)
10.1038/nbt1102-1082 / Nature Biotechnology / An open-source oligomicroarray standard for human and mouse by Wright (2002)
10.1038/nature08180 / Nature / Elite and stochastic models for induced pluripotent stem cell generation by Yamanaka (2009)
10.1128/AEM.72.1.733-744.2006 / Applied and Environmental Microbiology / Making all parts of the 16S rRNA of Escherichia coli accessible in situ to single DNA oligonucleotides by Yilmaz (2006)
10.1074/jbc.M604369200 / Journal of Biological Chemistry / Observed instability of T7 RNA polymerase elongation complexes can be dominated by collision-induced “bumping” by Zhou (2006)
10.1529/biophysj.105.071027 / Biophysical Journal / Fluorescence quenching by TEMPO: a sub-30 A single-molecule ruler by Zhu (2005)
{'key': 'bib66', 'first-page': '75', 'article-title': 'On the complexity of finite sequences', 'volume': 'IT-22', 'author': 'Ziv', 'year': '1976', 'journal-title': 'IEEE Transactions on Information Theory'} / IEEE Transactions on Information Theory / On the complexity of finite sequences by Ziv (1976)
10.1093/nar/gkg595 / Nucleic Acids Research / Mfold web server for nucleic acid folding and hybridization prediction by Zuker (2003)

Dates

Type	When
Created	11 years, 7 months ago (Jan. 28, 2014, 11:24 a.m.)
Deposited	1 year, 10 months ago (Oct. 11, 2023, 8:43 p.m.)
Indexed	1 month ago (July 30, 2025, 1:07 p.m.)
Issued	11 years, 7 months ago (Jan. 28, 2014)
Published	11 years, 7 months ago (Jan. 28, 2014)
Published Online	11 years, 7 months ago (Jan. 28, 2014)

Funders 2

Howard Hughes Medical Institute 10.13039/100000011
Region: Americas
pri (Research institutes and centers)
Labels2
1. Howard Hughes Medical Institute Inc
2. HHMI
Howard Hughes Medical Institute 10.13039/100000011
Region: Americas
pri (Research institutes and centers)
Labels2
1. Howard Hughes Medical Institute Inc
2. HHMI

BibTeX

@article{Zhang_2014, title={Single-molecule tracking of the transcription cycle by sub-second RNA detection}, volume={3}, ISSN={2050-084X}, url={http://dx.doi.org/10.7554/elife.01775}, DOI={10.7554/elife.01775}, journal={eLife}, publisher={eLife Sciences Publications, Ltd}, author={Zhang, Zhengjian and Revyakin, Andrey and Grimm, Jonathan B and Lavis, Luke D and Tjian, Robert}, year={2014}, month=jan }

JSON

{
  "indexed": {
    "date-parts": [
      [
        2025,
        7,
        30
      ]
    ],
    "date-time": "2025-07-30T17:07:47Z",
    "timestamp": 1753895267180,
    "version": "3.41.2"
  },
  "reference-count": 67,
  "publisher": "eLife Sciences Publications, Ltd",
  "license": [
    {
      "start": {
        "date-parts": [
          [
            2014,
            1,
            28
          ]
        ],
        "date-time": "2014-01-28T00:00:00Z",
        "timestamp": 1390867200000
      },
      "content-version": "vor",
      "delay-in-days": 0,
      "URL": "http://creativecommons.org/licenses/by/3.0/"
    },
    {
      "start": {
        "date-parts": [
          [
            2014,
            1,
            28
          ]
        ],
        "date-time": "2014-01-28T00:00:00Z",
        "timestamp": 1390867200000
      },
      "content-version": "am",
      "delay-in-days": 0,
      "URL": "http://creativecommons.org/licenses/by/3.0/"
    },
    {
      "start": {
        "date-parts": [
          [
            2014,
            1,
            28
          ]
        ],
        "date-time": "2014-01-28T00:00:00Z",
        "timestamp": 1390867200000
      },
      "content-version": "tdm",
      "delay-in-days": 0,
      "URL": "http://creativecommons.org/licenses/by/3.0/"
    },
    {
      "start": {
        "date-parts": [
          [
            2014,
            1,
            28
          ]
        ],
        "date-time": "2014-01-28T00:00:00Z",
        "timestamp": 1390867200000
      },
      "content-version": "vor",
      "delay-in-days": 0,
      "URL": "http://creativecommons.org/licenses/by/3.0/"
    },
    {
      "start": {
        "date-parts": [
          [
            2014,
            1,
            28
          ]
        ],
        "date-time": "2014-01-28T00:00:00Z",
        "timestamp": 1390867200000
      },
      "content-version": "am",
      "delay-in-days": 0,
      "URL": "http://creativecommons.org/licenses/by/3.0/"
    },
    {
      "start": {
        "date-parts": [
          [
            2014,
            1,
            28
          ]
        ],
        "date-time": "2014-01-28T00:00:00Z",
        "timestamp": 1390867200000
      },
      "content-version": "tdm",
      "delay-in-days": 0,
      "URL": "http://creativecommons.org/licenses/by/3.0/"
    }
  ],
  "funder": [
    {
      "DOI": "10.13039/100000011",
      "name": "Howard Hughes Medical Institute",
      "doi-asserted-by": "crossref",
      "id": [
        {
          "id": "10.13039/100000011",
          "id-type": "DOI",
          "asserted-by": "crossref"
        }
      ]
    },
    {
      "DOI": "10.13039/100000011",
      "name": "Howard Hughes Medical Institute",
      "doi-asserted-by": "publisher",
      "id": [
        {
          "id": "10.13039/100000011",
          "id-type": "DOI",
          "asserted-by": "publisher"
        }
      ]
    }
  ],
  "content-domain": {
    "domain": [
      "www.elifesciences.org"
    ],
    "crossmark-restriction": false
  },
  "abstract": "<jats:p>Transcription is an inherently stochastic, noisy, and multi-step process, in which fluctuations at every step can cause variations in RNA synthesis, and affect physiology and differentiation decisions in otherwise identical cells. However, it has been an experimental challenge to directly link the stochastic events at the promoter to transcript production. Here we established a fast fluorescence in situ hybridization (fastFISH) method that takes advantage of intrinsically unstructured nucleic acid sequences to achieve exceptionally fast rates of specific hybridization (\u223c10e7 M\u22121s\u22121), and allows deterministic detection of single nascent transcripts. Using a prototypical RNA polymerase, we demonstrated the use of fastFISH to measure the kinetic rates of promoter escape, elongation, and termination in one assay at the single-molecule level, at sub-second temporal resolution. The principles of fastFISH design can be used to study stochasticity in gene regulation, to select targets for gene silencing, and to design nucleic acid nanostructures.</jats:p>",
  "DOI": "10.7554/elife.01775",
  "type": "journal-article",
  "created": {
    "date-parts": [
      [
        2014,
        1,
        28
      ]
    ],
    "date-time": "2014-01-28T16:24:12Z",
    "timestamp": 1390926252000
  },
  "source": "Crossref",
  "is-referenced-by-count": 70,
  "title": "Single-molecule tracking of the transcription cycle by sub-second RNA detection",
  "prefix": "10.7554",
  "volume": "3",
  "author": [
    {
      "given": "Zhengjian",
      "family": "Zhang",
      "sequence": "first",
      "affiliation": [
        {
          "name": "Janelia Farm Research Campus, Howard Hughes Medical Institute, Ashburn, United States"
        }
      ]
    },
    {
      "given": "Andrey",
      "family": "Revyakin",
      "sequence": "additional",
      "affiliation": [
        {
          "name": "Janelia Farm Research Campus, Howard Hughes Medical Institute, Ashburn, United States"
        }
      ]
    },
    {
      "given": "Jonathan B",
      "family": "Grimm",
      "sequence": "additional",
      "affiliation": [
        {
          "name": "Janelia Farm Research Campus, Howard Hughes Medical Institute, Ashburn, United States"
        }
      ]
    },
    {
      "given": "Luke D",
      "family": "Lavis",
      "sequence": "additional",
      "affiliation": [
        {
          "name": "Janelia Farm Research Campus, Howard Hughes Medical Institute, Ashburn, United States"
        }
      ]
    },
    {
      "given": "Robert",
      "family": "Tjian",
      "sequence": "additional",
      "affiliation": [
        {
          "name": "Janelia Farm Research Campus, Howard Hughes Medical Institute, Ashburn, United States"
        },
        {
          "name": "Li Ka Shing Center for Biomedical and Health Sciences, University of California, Berkeley, Berkeley, United States"
        },
        {
          "name": "Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States"
        }
      ]
    }
  ],
  "member": "4374",
  "published-online": {
    "date-parts": [
      [
        2014,
        1,
        28
      ]
    ]
  },
  "reference": [
    {
      "key": "bib1",
      "doi-asserted-by": "publisher",
      "first-page": "271",
      "DOI": "10.1016/0022-2836(88)90457-3",
      "article-title": "Transcription termination in Escherichia coli. Measurement of the rate of enzyme release from Rho-independent terminators",
      "volume": "202",
      "author": "Arndt",
      "year": "1988",
      "journal-title": "Journal of Molecular Biology"
    },
    {
      "key": "bib2",
      "doi-asserted-by": "publisher",
      "first-page": "343",
      "DOI": "10.1146/annurev.biophys.35.010406.150153",
      "article-title": "Single-molecule analysis of RNA polymerase transcription",
      "volume": "35",
      "author": "Bai",
      "year": "2006",
      "journal-title": "Annual Review of Biophysics and Biomolecular Structure"
    },
    {
      "key": "bib3",
      "doi-asserted-by": "publisher",
      "first-page": "12893",
      "DOI": "10.1073/pnas.0403884101",
      "article-title": "tRNA dynamics on the ribosome during translation",
      "volume": "101",
      "author": "Blanchard",
      "year": "2004",
      "journal-title": "Proceedings of the National Academy of Sciences of the United States of America"
    },
    {
      "key": "bib4",
      "doi-asserted-by": "crossref",
      "first-page": "25120",
      "DOI": "10.1016/S0021-9258(17)31506-5",
      "article-title": "Characterization of a set of T7 RNA polymerase active site mutants",
      "volume": "269",
      "author": "Bonner",
      "year": "1994",
      "journal-title": "Journal of Biological Chemistry"
    },
    {
      "key": "bib5",
      "doi-asserted-by": "publisher",
      "first-page": "591",
      "DOI": "10.1126/science.1218716",
      "article-title": "Opening and closing of the bacterial RNA polymerase clamp",
      "volume": "337",
      "author": "Chakraborty",
      "year": "2012",
      "journal-title": "Science"
    },
    {
      "key": "bib6",
      "doi-asserted-by": "publisher",
      "first-page": "2243",
      "DOI": "10.1016/S0006-3495(95)80095-0",
      "article-title": "The biophysics of DNA hybridization with immobilized oligonucleotide probes",
      "volume": "69",
      "author": "Chan",
      "year": "1995",
      "journal-title": "Biophysical Journal"
    },
    {
      "key": "bib7",
      "doi-asserted-by": "publisher",
      "first-page": "1431",
      "DOI": "10.1016/j.cell.2012.06.006",
      "article-title": "A movie of RNA polymerase ii transcription",
      "volume": "149",
      "author": "Cheung",
      "year": "2012",
      "journal-title": "Cell"
    },
    {
      "key": "bib8",
      "doi-asserted-by": "crossref",
      "first-page": "3019",
      "DOI": "10.1128/JB.180.12.3019-3025.1998",
      "article-title": "RNA polymerase-promoter interactions: the comings and goings of RNA Polymerase",
      "volume": "180",
      "author": "DeHaseth",
      "year": "1998",
      "journal-title": "Journal of Bacteriology"
    },
    {
      "key": "bib9",
      "doi-asserted-by": "publisher",
      "first-page": "248102",
      "DOI": "10.1103/PhysRevLett.89.248102",
      "article-title": "Stretching single stranded DNA, a model polyelectrolyte",
      "volume": "89",
      "author": "Dessinges",
      "year": "2002",
      "journal-title": "Physical Review Letters"
    },
    {
      "key": "bib10",
      "doi-asserted-by": "publisher",
      "first-page": "801",
      "DOI": "10.1126/science.1083219",
      "article-title": "Cooperation between RNA polymerase molecules in transcription elongation",
      "volume": "300",
      "author": "Epshtein",
      "year": "2003",
      "journal-title": "Science"
    },
    {
      "key": "bib11",
      "doi-asserted-by": "publisher",
      "first-page": "1023",
      "DOI": "10.1529/biophysj.106.084004",
      "article-title": "Viewing dynamic assembly of molecular complexes by multi-wavelength single-molecule fluorescence",
      "volume": "91",
      "author": "Friedman",
      "year": "2006",
      "journal-title": "Biophysical Journal"
    },
    {
      "key": "bib12",
      "doi-asserted-by": "publisher",
      "first-page": "679",
      "DOI": "10.1016/j.cell.2012.01.018",
      "article-title": "Mechanism of transcription initiation at an activator-dependent promoter defined by single-molecule observation",
      "volume": "148",
      "author": "Friedman",
      "year": "2012",
      "journal-title": "Cell"
    },
    {
      "key": "bib13",
      "doi-asserted-by": "publisher",
      "first-page": "9740",
      "DOI": "10.1073/pnas.1300221110",
      "article-title": "RNA polymerase approaches its promoter without long-range sliding along DNA",
      "volume": "110",
      "author": "Friedman",
      "year": "2013",
      "journal-title": "Proceedings of the National Academy of Sciences of the United States of America"
    },
    {
      "key": "bib14",
      "doi-asserted-by": "publisher",
      "first-page": "820",
      "DOI": "10.1038/nature05701",
      "article-title": "Backtracking determines the force sensitivity of RNAP II in a factor-dependent manner",
      "volume": "446",
      "author": "Galburt",
      "year": "2007",
      "journal-title": "Nature"
    },
    {
      "key": "bib15",
      "doi-asserted-by": "publisher",
      "first-page": "3370",
      "DOI": "10.1093/nar/gkl422",
      "article-title": "Secondary structure effects on DNA hybridization kinetics: a solution versus surface comparison",
      "volume": "34",
      "author": "Gao",
      "year": "2006",
      "journal-title": "Nucleic Acids Research"
    },
    {
      "key": "bib16",
      "doi-asserted-by": "crossref",
      "first-page": "2858",
      "DOI": "10.1016/S0021-9258(19)42709-9",
      "article-title": "Characterization of T7-specific ribonucleic acid polymerase: IV. Resolution of the major in vitro transcripts by gel electrophoresis",
      "volume": "249",
      "author": "Golomb",
      "year": "1974",
      "journal-title": "Journal of Biological Chemistry"
    },
    {
      "key": "bib17",
      "doi-asserted-by": "publisher",
      "first-page": "3314",
      "DOI": "10.1016/S0006-3495(02)75672-5",
      "article-title": "Magnetic tweezers: micromanipulation and force measurement at the molecular level",
      "volume": "82",
      "author": "Gosse",
      "year": "2002",
      "journal-title": "Biophysical Journal"
    },
    {
      "key": "bib18",
      "doi-asserted-by": "publisher",
      "first-page": "W70",
      "DOI": "10.1093/nar/gkn188",
      "article-title": "The Vienna RNA websuite",
      "volume": "36",
      "author": "Gruber",
      "year": "2008",
      "journal-title": "Nucleic Acids Research"
    },
    {
      "key": "bib19",
      "doi-asserted-by": "publisher",
      "first-page": "633",
      "DOI": "10.1016/j.cell.2011.07.026",
      "article-title": "Stochastic state transitions give rise to phenotypic equilibrium in populations of cancer cells",
      "volume": "146",
      "author": "Gupta",
      "year": "2011",
      "journal-title": "Cell"
    },
    {
      "key": "bib20",
      "doi-asserted-by": "publisher",
      "first-page": "741",
      "DOI": "10.1016/j.molcel.2009.09.005",
      "article-title": "Transcription dynamics",
      "volume": "35",
      "author": "Hager",
      "year": "2009",
      "journal-title": "Molecular Cell"
    },
    {
      "key": "bib21",
      "doi-asserted-by": "publisher",
      "first-page": "993",
      "DOI": "10.1146/annurev.biochem.77.061906.092014",
      "article-title": "Super resolution fluorescence microscopy",
      "volume": "78",
      "author": "Huang",
      "year": "2010",
      "journal-title": "Annual Review of Biochemistry"
    },
    {
      "key": "bib22",
      "doi-asserted-by": "publisher",
      "first-page": "347",
      "DOI": "10.1006/jmbi.2000.4150",
      "article-title": "T7 RNA polymerase elongation complex structure and movement",
      "volume": "303",
      "author": "Huang",
      "year": "2000",
      "journal-title": "Journal of Molecular Biology"
    },
    {
      "key": "bib23",
      "doi-asserted-by": "publisher",
      "first-page": "2290",
      "DOI": "10.1074/jbc.270.5.2290",
      "article-title": "RNA polymerase II ternary complexes may become arrested after transcribing to within 10 bases of the end of linear templates",
      "volume": "270",
      "author": "Izban",
      "year": "1995",
      "journal-title": "Journal of Biological Chemistry"
    },
    {
      "key": "bib24",
      "doi-asserted-by": "publisher",
      "first-page": "6950",
      "DOI": "10.1021/ja025678o",
      "article-title": "Intramolecular dimers: a new strategy to fluorescence quenching in dual-labeled oligonucleotide probes",
      "volume": "124",
      "author": "Johansson",
      "year": "2002",
      "journal-title": "Journal of the American Chemical Society"
    },
    {
      "key": "bib25",
      "doi-asserted-by": "publisher",
      "first-page": "1144",
      "DOI": "10.1126/science.1131399",
      "article-title": "Initial transcription by RNA polymerase proceeds through a DNA-scrunching mechanism",
      "volume": "314",
      "author": "Kapanidis",
      "year": "2006",
      "journal-title": "Science"
    },
    {
      "key": "bib26",
      "doi-asserted-by": "publisher",
      "first-page": "842",
      "DOI": "10.1103/PhysRevA.36.842",
      "article-title": "Easily calculable measure for the complexity of spatiotemporal patterns",
      "volume": "36",
      "author": "Kaspar",
      "year": "1987",
      "journal-title": "Physical Review A"
    },
    {
      "key": "bib27",
      "doi-asserted-by": "publisher",
      "first-page": "1097",
      "DOI": "10.1021/ja0041324",
      "article-title": "Hybridization of DNA and PNA molecular beacons to single-stranded and double-stranded DNA targets",
      "volume": "124",
      "author": "Kuhn",
      "year": "2002",
      "journal-title": "Journal of the American Chemical Society"
    },
    {
      "key": "bib28",
      "doi-asserted-by": "publisher",
      "first-page": "10805",
      "DOI": "10.1021/ja016310e",
      "article-title": "Effect of secondary structure on the thermodynamics and kinetics of PNA hybridization to DNA hairpins",
      "volume": "123",
      "author": "Kushon",
      "year": "2001",
      "journal-title": "Journal of the American Chemical Society"
    },
    {
      "key": "bib29",
      "doi-asserted-by": "publisher",
      "first-page": "12055",
      "DOI": "10.1021/bi00163a013",
      "article-title": "Implication of RNA structure on antisense oligonucleotide hybridization kinetics",
      "volume": "31",
      "author": "Lima",
      "year": "1992",
      "journal-title": "Biochemistry"
    },
    {
      "key": "bib30",
      "doi-asserted-by": "publisher",
      "first-page": "313",
      "DOI": "10.1038/embor.2012.31",
      "article-title": "Transcription goes digital",
      "volume": "13",
      "author": "Lionnet",
      "year": "2012",
      "journal-title": "EMBO Reports"
    },
    {
      "key": "bib31",
      "doi-asserted-by": "publisher",
      "first-page": "1580",
      "DOI": "10.1126/science.1235441",
      "article-title": "Transcription under torsion",
      "volume": "340",
      "author": "Ma",
      "year": "2013",
      "journal-title": "Science"
    },
    {
      "key": "bib32",
      "doi-asserted-by": "crossref",
      "first-page": "e122",
      "DOI": "10.1093/nar/gnf121",
      "article-title": "Efficiencies of fluorescence resonance energy transfer and contact-mediated quenching in oligonucleotide probes",
      "volume": "30",
      "author": "Marras",
      "year": "2002",
      "journal-title": "Nucleic Acids Research"
    },
    {
      "key": "bib33",
      "doi-asserted-by": "publisher",
      "first-page": "2690",
      "DOI": "10.1021/bi00384a006",
      "article-title": "Kinetic analysis of T7 RNA polymerase-promoter interactions with small synthetic promoters",
      "volume": "26",
      "author": "Martin",
      "year": "1987",
      "journal-title": "Biochemistry"
    },
    {
      "key": "bib34",
      "doi-asserted-by": "publisher",
      "first-page": "4281",
      "DOI": "10.1021/bi00067a017",
      "article-title": "Kinetic analysis of T7 RNA polymerase transcription initiation from promoters containing single-stranded regions",
      "volume": "32",
      "author": "Maslak",
      "year": "1993",
      "journal-title": "Biochemistry"
    },
    {
      "key": "bib35",
      "doi-asserted-by": "publisher",
      "first-page": "8783",
      "DOI": "10.1093/nar/15.21.8783",
      "article-title": "Oligoribonucleotide synthesis using T7 RNA polymerase and synthetic DNA templates",
      "volume": "15",
      "author": "Milligan",
      "year": "1987",
      "journal-title": "Nucleic Acids Research"
    },
    {
      "key": "bib35a",
      "doi-asserted-by": "publisher",
      "first-page": "105",
      "DOI": "10.1021/bc00020a001",
      "article-title": "Cyanine dye labeling reagents: sulfoindocyanine succinimidyl esters",
      "volume": "4",
      "author": "Mujumdar",
      "year": "1993",
      "journal-title": "Bioconjugate Chemistry"
    },
    {
      "key": "bib36",
      "doi-asserted-by": "publisher",
      "first-page": "31",
      "DOI": "10.1016/S0959-440X(02)00005-2",
      "article-title": "Bacterial RNA polymerases: the wholo story",
      "volume": "13",
      "author": "Murakami",
      "year": "2003",
      "journal-title": "Current Opinion in Structural Biology"
    },
    {
      "key": "bib37",
      "doi-asserted-by": "publisher",
      "first-page": "437",
      "DOI": "10.1016/S0092-8674(03)00845-6",
      "article-title": "Ubiquitous transcriptional pausing is independent of RNA polymerase backtracking",
      "volume": "115",
      "author": "Neuman",
      "year": "2003",
      "journal-title": "Cell"
    },
    {
      "key": "bib38",
      "doi-asserted-by": "crossref",
      "first-page": "2844",
      "DOI": "10.1182/blood.V81.11.2844.2844",
      "article-title": "Differentiation and proliferation of hematopoietic stem cells",
      "volume": "81",
      "author": "Ogawa",
      "year": "1993",
      "journal-title": "Blood"
    },
    {
      "key": "bib39",
      "doi-asserted-by": "publisher",
      "first-page": "W628",
      "DOI": "10.1093/nar/gkh466",
      "article-title": "Complexity: an internet resource for analysis of DNA sequence complexity",
      "volume": "32",
      "author": "Orlov",
      "year": "2004",
      "journal-title": "Nucleic Acids Research"
    },
    {
      "key": "bib40",
      "doi-asserted-by": "publisher",
      "first-page": "153",
      "DOI": "10.1002/9780470559277.ch110148",
      "article-title": "Copper-catalyzed azide-alkyne click chemistry for bioconjugation",
      "volume": "3",
      "author": "Presolski",
      "year": "2011",
      "journal-title": "Current Protocols in Chemical Biology"
    },
    {
      "key": "bib41",
      "doi-asserted-by": "publisher",
      "first-page": "255",
      "DOI": "10.1146/annurev.biophys.37.032807.125928",
      "article-title": "Single-molecule approaches to stochastic gene expression",
      "volume": "38",
      "author": "Raj",
      "year": "2009",
      "journal-title": "Annual Review of Biophysics"
    },
    {
      "key": "bib42",
      "doi-asserted-by": "publisher",
      "first-page": "3147",
      "DOI": "10.1073/pnas.90.8.3147",
      "article-title": "T7 RNA polymerase mutants with altered promoter specificities",
      "volume": "90",
      "author": "Raskin",
      "year": "1993",
      "journal-title": "Proceedings of the National Academy of Sciences of the United States of America"
    },
    {
      "key": "bib43",
      "doi-asserted-by": "publisher",
      "first-page": "1139",
      "DOI": "10.1126/science.1131398",
      "article-title": "Abortive initiation and productive initiation by RNA polymerase involve DNA scrunching",
      "volume": "314",
      "author": "Revyakin",
      "year": "2006",
      "journal-title": "Science"
    },
    {
      "key": "bib44",
      "doi-asserted-by": "publisher",
      "first-page": "1691",
      "DOI": "10.1101/gad.194936.112",
      "article-title": "Transcription initiation by human RNA polymerase II visualized at single-molecule resolution",
      "volume": "26",
      "author": "Revyakin",
      "year": "2012",
      "journal-title": "Genes & Development"
    },
    {
      "key": "bib45",
      "doi-asserted-by": "publisher",
      "first-page": "875",
      "DOI": "10.1016/j.jmb.2012.12.015",
      "article-title": "The transcription bubble of the RNA polymerase-promoter open complex exhibits conformational heterogeneity and millisecond-scale dynamics: implications for transcription start-site selection",
      "volume": "425",
      "author": "Robb",
      "year": "2013",
      "journal-title": "Journal of Molecular Biology"
    },
    {
      "key": "bib46",
      "doi-asserted-by": "publisher",
      "first-page": "297",
      "DOI": "10.1038/nature04586",
      "article-title": "Folding DNA to create nanoscale shapes and patterns",
      "volume": "440",
      "author": "Rothemund",
      "year": "2006",
      "journal-title": "Nature"
    },
    {
      "key": "bib47",
      "doi-asserted-by": "crossref",
      "DOI": "10.1002/9780470316849",
      "volume-title": "Multivariate density estimation: theory, practice, and visualization (Wiley series in probability and statistics)",
      "author": "Scott",
      "year": "1992"
    },
    {
      "key": "bib48",
      "doi-asserted-by": "publisher",
      "first-page": "684",
      "DOI": "10.1038/nature02191",
      "article-title": "Backtracking by single RNA polymerase molecules observed at near-base-pair resolution",
      "volume": "426",
      "author": "Shaevitz",
      "year": "2003",
      "journal-title": "Nature"
    },
    {
      "key": "bib49",
      "doi-asserted-by": "publisher",
      "first-page": "3239",
      "DOI": "10.1074/jbc.M310471200",
      "article-title": "Promoter binding, initiation, and elongation by bacteriophage T7 RNA polymerase. A single-molecule view of the transcription cycle",
      "volume": "279",
      "author": "Skinner",
      "year": "2004",
      "journal-title": "Journal of Biological Chemistry"
    },
    {
      "key": "bib50",
      "doi-asserted-by": "publisher",
      "first-page": "795",
      "DOI": "10.1126/science.271.5250.795",
      "article-title": "Overstretching B-DNA: the elastic response of individual double-stranded and single-stranded DNA molecules",
      "volume": "271",
      "author": "Smith",
      "year": "1996",
      "journal-title": "Science"
    },
    {
      "key": "bib52",
      "doi-asserted-by": "publisher",
      "first-page": "37292",
      "DOI": "10.1074/jbc.M201600200",
      "article-title": "The +2 NTP binding drives open complex formation in T7 RNA polymerase",
      "volume": "277",
      "author": "Stano",
      "year": "2002",
      "journal-title": "Journal of Biological Chemistry"
    },
    {
      "key": "bib53",
      "doi-asserted-by": "publisher",
      "first-page": "4947",
      "DOI": "10.1021/bi052292s",
      "article-title": "Rapid binding of T7 RNA polymerase is followed by simultaneous bending and opening of the promoter DNA",
      "volume": "45",
      "author": "Tang",
      "year": "2006",
      "journal-title": "Biochemistry"
    },
    {
      "key": "bib54",
      "doi-asserted-by": "publisher",
      "first-page": "22175",
      "DOI": "10.1073/pnas.0906979106",
      "article-title": "Real-time observation of the transition from transcription initiation to elongation of the RNA polymerase",
      "volume": "106",
      "author": "Tang",
      "year": "2009",
      "journal-title": "Proceedings of the National Academy of Sciences of the United States of America"
    },
    {
      "key": "bib55",
      "doi-asserted-by": "publisher",
      "first-page": "2423",
      "DOI": "10.1529/biophysj.107.125096",
      "article-title": "T7 RNA polymerase studied by force measurements varying cofactor concentration",
      "volume": "95",
      "author": "Thomen",
      "year": "2008",
      "journal-title": "Biophysical Journal"
    },
    {
      "key": "bib56",
      "doi-asserted-by": "publisher",
      "first-page": "1319",
      "DOI": "10.1093/nar/gkg212",
      "article-title": "Hybridization kinetics and thermodynamics of molecular beacons",
      "volume": "31",
      "author": "Tsourkas",
      "year": "2003",
      "journal-title": "Nucleic Acids Research"
    },
    {
      "key": "bib57",
      "doi-asserted-by": "publisher",
      "first-page": "174",
      "DOI": "10.1038/nsmb.2472",
      "article-title": "The promoter-search mechanism of Escherichia coli RNA polymerase is dominated by three-dimensional diffusion",
      "volume": "20",
      "author": "Wang",
      "year": "2012",
      "journal-title": "Nature Structure Molecular Biology"
    },
    {
      "key": "bib58",
      "doi-asserted-by": "publisher",
      "first-page": "167",
      "DOI": "10.1089/oli.2004.14.167",
      "article-title": "Computational identification of antisense oligonucleotides that rapidly hybridize to RNA",
      "volume": "14",
      "author": "Wang",
      "year": "2004",
      "journal-title": "Oligonucleotides"
    },
    {
      "key": "bib59",
      "doi-asserted-by": "publisher",
      "first-page": "902",
      "DOI": "10.1126/science.282.5390.902",
      "article-title": "Force and velocity measured for single molecules of RNA polymerase",
      "volume": "282",
      "author": "Wang",
      "year": "1998",
      "journal-title": "Science"
    },
    {
      "key": "bib60",
      "doi-asserted-by": "publisher",
      "first-page": "1676",
      "DOI": "10.1126/science.283.5408.1676",
      "article-title": "Fluorescence spectroscopy of single biomolecules",
      "volume": "283",
      "author": "Weiss",
      "year": "1999",
      "journal-title": "Science"
    },
    {
      "key": "bib61",
      "doi-asserted-by": "publisher",
      "first-page": "1082",
      "DOI": "10.1038/nbt1102-1082",
      "article-title": "An open-source oligomicroarray standard for human and mouse",
      "volume": "20",
      "author": "Wright",
      "year": "2002",
      "journal-title": "Nature Biotechnology"
    },
    {
      "key": "bib62",
      "doi-asserted-by": "publisher",
      "first-page": "49",
      "DOI": "10.1038/nature08180",
      "article-title": "Elite and stochastic models for induced pluripotent stem cell generation",
      "volume": "460",
      "author": "Yamanaka",
      "year": "2009",
      "journal-title": "Nature"
    },
    {
      "key": "bib63",
      "doi-asserted-by": "publisher",
      "first-page": "733",
      "DOI": "10.1128/AEM.72.1.733-744.2006",
      "article-title": "Making all parts of the 16S rRNA of Escherichia coli accessible in situ to single DNA oligonucleotides",
      "volume": "72",
      "author": "Yilmaz",
      "year": "2006",
      "journal-title": "Applied and Environmental Microbiology"
    },
    {
      "key": "bib64",
      "doi-asserted-by": "publisher",
      "first-page": "24441",
      "DOI": "10.1074/jbc.M604369200",
      "article-title": "Observed instability of T7 RNA polymerase elongation complexes can be dominated by collision-induced \u201cbumping\u201d",
      "volume": "281",
      "author": "Zhou",
      "year": "2006",
      "journal-title": "Journal of Biological Chemistry"
    },
    {
      "key": "bib65",
      "doi-asserted-by": "publisher",
      "first-page": "L37",
      "DOI": "10.1529/biophysj.105.071027",
      "article-title": "Fluorescence quenching by TEMPO: a sub-30 A single-molecule ruler",
      "volume": "89",
      "author": "Zhu",
      "year": "2005",
      "journal-title": "Biophysical Journal"
    },
    {
      "key": "bib66",
      "first-page": "75",
      "article-title": "On the complexity of finite sequences",
      "volume": "IT-22",
      "author": "Ziv",
      "year": "1976",
      "journal-title": "IEEE Transactions on Information Theory"
    },
    {
      "key": "bib67",
      "doi-asserted-by": "publisher",
      "first-page": "3406",
      "DOI": "10.1093/nar/gkg595",
      "article-title": "Mfold web server for nucleic acid folding and hybridization prediction",
      "volume": "31",
      "author": "Zuker",
      "year": "2003",
      "journal-title": "Nucleic Acids Research"
    }
  ],
  "container-title": "eLife",
  "original-title": [],
  "language": "en",
  "link": [
    {
      "URL": "https://cdn.elifesciences.org/articles/01775/elife-01775-v1.pdf",
      "content-type": "application/pdf",
      "content-version": "vor",
      "intended-application": "text-mining"
    },
    {
      "URL": "https://cdn.elifesciences.org/articles/01775/elife-01775-v1.xml",
      "content-type": "application/xml",
      "content-version": "vor",
      "intended-application": "text-mining"
    },
    {
      "URL": "https://elifesciences.org/articles/01775",
      "content-type": "unspecified",
      "content-version": "vor",
      "intended-application": "similarity-checking"
    }
  ],
  "deposited": {
    "date-parts": [
      [
        2023,
        10,
        12
      ]
    ],
    "date-time": "2023-10-12T00:43:25Z",
    "timestamp": 1697071405000
  },
  "score": 1,
  "resource": {
    "primary": {
      "URL": "https://elifesciences.org/articles/01775"
    }
  },
  "subtitle": [],
  "short-title": [],
  "issued": {
    "date-parts": [
      [
        2014,
        1,
        28
      ]
    ]
  },
  "references-count": 67,
  "alternative-id": [
    "10.7554/eLife.01775"
  ],
  "URL": "http://dx.doi.org/10.7554/eLife.01775",
  "relation": {
    "has-review": [
      {
        "id-type": "doi",
        "id": "10.7554/eLife.01775.017",
        "asserted-by": "object"
      },
      {
        "id-type": "doi",
        "id": "10.7554/eLife.01775.016",
        "asserted-by": "object"
      }
    ]
  },
  "ISSN": [
    "2050-084X"
  ],
  "subject": [],
  "published": {
    "date-parts": [
      [
        2014,
        1,
        28
      ]
    ]
  },
  "assertion": [
    {
      "value": "2013-10-24",
      "order": 0,
      "name": "received",
      "label": "Received",
      "group": {
        "name": "publication_history",
        "label": "Publication History"
      }
    },
    {
      "value": "2013-12-06",
      "order": 1,
      "name": "accepted",
      "label": "Accepted",
      "group": {
        "name": "publication_history",
        "label": "Publication History"
      }
    },
    {
      "value": "2014-01-28",
      "order": 2,
      "name": "published",
      "label": "Published",
      "group": {
        "name": "publication_history",
        "label": "Publication History"
      }
    }
  ],
  "article-number": "e01775"
}