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Active Ribosome Profiling With RiboLace

56 Pages Posted: 12 Apr 2018 Sneak Peek Status: Published

See all articles by Massimiliano Clamer

Massimiliano Clamer

University of Trento - Centre for Integrative Biology (CIBIO)

Toma Tebaldi

University of Trento - Centre for Integrative Biology (CIBIO)

Fabio Lauria

Italian National Research Council (CNR) - Institute of Biophysics, Trento

Paola Bernabò

Italian National Research Council (CNR) - Institute of Biophysics, Trento

Rodolfo F. Gómez-Biagi

Fondazione Bruno Kessler

Elena Perenthaler

Italian National Research Council (CNR) - Institute of Biophysics, Trento

Daniele Gubert

Italian National Research Council (CNR) - Institute of Biophysics, Trento

Laura Pasquardini

Fondazione Bruno Kessler

Graziano Guella

University of Trento - Centre for Integrative Biology (CIBIO)

Ewout J. N. Groen

University of Edinburgh - Biomedical Sciences

Thomas H. Gillingwater

University of Edinburgh - Biomedical Sciences

Alessandro Quattrone

University of Trento - Centre for Integrative Biology (CIBIO)

Gabriella Viero

Italian National Research Council (CNR) - Institute of Biophysics, Trento

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Abstract

Ribosome profiling, or Ribo-Seq, is based on large-scale sequencing of RNA fragments protected from nuclease digestion by ribosomes. Thanks to its unique ability to provide positional information about ribosomes flowing along transcripts, this method can be used to shed light on mechanistic aspects of translation. However, current Ribo-Seq approaches lack the ability to distinguish between fragments protected either by ribosomes in active translation or by inactive ribosomes. To overcome this significant limitation, we developed RiboLace: a novel method based on an original puromycin-containing molecule capable of isolating active ribosomes by means of an antibody-free and tag-free pulldown approach. RiboLace is fast, works reliably with low amounts of input material, and can be easily and rapidly applied both in vitro and in vivo, thereby generating a global snapshot of active ribosome footprints at single nucleotide resolution.

Suggested Citation

Clamer, Massimiliano and Tebaldi, Toma and Lauria, Fabio and Bernabò, Paola and Gómez-Biagi, Rodolfo F. and Perenthaler, Elena and Gubert, Daniele and Pasquardini, Laura and Guella, Graziano and Groen, Ewout J. N. and Gillingwater, Thomas H. and Quattrone, Alessandro and Viero, Gabriella, Active Ribosome Profiling With RiboLace (2018). Available at SSRN: https://ssrn.com/abstract=3155822 or http://dx.doi.org/10.2139/ssrn.3155822
This is a paper under consideration at Cell Press and has not been peer-reviewed.

Massimiliano Clamer

University of Trento - Centre for Integrative Biology (CIBIO)

via Sommarive n. 9, Povo (TN)
Povo, Trentino 38123
Italy

Toma Tebaldi

University of Trento - Centre for Integrative Biology (CIBIO)

via Sommarive n. 9, Povo (TN)
Povo, Trentino 38123
Italy

Fabio Lauria

Italian National Research Council (CNR) - Institute of Biophysics, Trento

Via alla Cascata, 56
Trento, Trentino 38123
Italy

Paola Bernabò

Italian National Research Council (CNR) - Institute of Biophysics, Trento

Via alla Cascata, 56
Trento, Trentino 38123
Italy

Rodolfo F. Gómez-Biagi

Fondazione Bruno Kessler

Via Sommarive 18
Povo
Trento, 38123
Italy

Elena Perenthaler

Italian National Research Council (CNR) - Institute of Biophysics, Trento

Via alla Cascata, 56
Trento, Trentino 38123
Italy

Daniele Gubert

Italian National Research Council (CNR) - Institute of Biophysics, Trento

Via alla Cascata, 56
Trento, Trentino 38123
Italy

Laura Pasquardini

Fondazione Bruno Kessler

Via Sommarive 18
Povo
Trento, 38123
Italy

Graziano Guella

University of Trento - Centre for Integrative Biology (CIBIO)

via Sommarive n. 9, Povo (TN)
Povo, Trentino 38123
Italy

Ewout J. N. Groen

University of Edinburgh - Biomedical Sciences

Edinburgh, Scotland EH8 9JY
United Kingdom

Thomas H. Gillingwater

University of Edinburgh - Biomedical Sciences

Edinburgh, Scotland EH8 9JY
United Kingdom

Alessandro Quattrone

University of Trento - Centre for Integrative Biology (CIBIO)

via Sommarive n. 9, Povo (TN)
Povo, Trentino 38123
Italy

Gabriella Viero (Contact Author)

Italian National Research Council (CNR) - Institute of Biophysics, Trento ( email )

Via alla Cascata, 56
Trento, Trentino 38123
Italy

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