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Method Article
Kostas D. Mathiopoulos
Laboratory of Molecular Biology and Genomics, Department of Biochemistry & Biotechnology, University of Thessaly, Larissa, Greece
Corresponding Author
Jiannis Ragoussis
McGill Genome Centre, Department of Human Genetics, McGill University, Montreal, Canada Department of Bioengineering, McGill University, Montréal, Québec, Canada
Corresponding Author
License:
This work is licensed under a CC BY 4.0 License. Read Full License
The Oxford Nanopore Technologies’ long-read RNA sequencing (RNAseq) platform can yield full length transcripts that can be very instrumental in improving the characterization of non-model organisms. The resolution of RNAseq can be increased by addition of external RNA molecules of known concentration such as ERCC in order to obtain absolute gene expression quantification. This protocol details the procedure to use ONT long-read RNAseq with addition of ERCC external RNAs. This protocol can be used with total RNA extracted from any source for which transcripts with a poly(A) tail at the 3’ end are the target. The protocol should be possible to complete in a single day (12 hours) followed by sequencing which takes another 48 hours.
Keywords
Nanopore,RNA sequencing,ERCC
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CURRENT STATUS: Posted
Version 1
Posted 10 May, 2021
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Molecular biology
Associated Publications
Nanopore long-read RNA-seq and absolute quantification delineate transcription dynamics in early embryo development of an insect pest
by Anthony Bayega, Spyros Oikonomopoulos, Maria-Eleni Gregoriou, +5
Scientific Reports (12 April, 2021)
Current and Future Methods for mRNA Analysis: A Drive Toward Single Molecule Sequencing
by Anthony Bayega, Somayyeh Fahiminiya, Spyros Oikonomopoulos, +1
Methods in Molecular Biology (16 May, 2018)
Transcript Profiling Using Long-Read Sequencing Technologies
by Anthony Bayega, Yu Chang Wang, Spyros Oikonomopoulos, +3
Methods in Molecular Biology (16 May, 2018)
An open repository of community-contributed protocols sponsored by Nature Research.
Learn more about Protocol Exchange
Method Article
Kostas D. Mathiopoulos
Laboratory of Molecular Biology and Genomics, Department of Biochemistry & Biotechnology, University of Thessaly, Larissa, Greece
Corresponding Author
Jiannis Ragoussis
McGill Genome Centre, Department of Human Genetics, McGill University, Montreal, Canada Department of Bioengineering, McGill University, Montréal, Québec, Canada
Corresponding Author
Version History
CURRENT STATUS: Posted
Version 1
Posted 10 May, 2021
No community comments so far
Metrics
Comments: 0
HTML Views: ...
Subject Areas
Molecular biology
License:
This work is licensed under a CC BY 4.0 License. Read Full License
The Oxford Nanopore Technologies’ long-read RNA sequencing (RNAseq) platform can yield full length transcripts that can be very instrumental in improving the characterization of non-model organisms. The resolution of RNAseq can be increased by addition of external RNA molecules of known concentration such as ERCC in order to obtain absolute gene expression quantification. This protocol details the procedure to use ONT long-read RNAseq with addition of ERCC external RNAs. This protocol can be used with total RNA extracted from any source for which transcripts with a poly(A) tail at the 3’ end are the target. The protocol should be possible to complete in a single day (12 hours) followed by sequencing which takes another 48 hours.
Loading...
Associated Publications
Nanopore long-read RNA-seq and absolute quantification delineate transcription dynamics in early embryo development of an insect pest
by Anthony Bayega, Spyros Oikonomopoulos, Maria-Eleni Gregoriou, +5
Scientific Reports (12 April, 2021)
Current and Future Methods for mRNA Analysis: A Drive Toward Single Molecule Sequencing
by Anthony Bayega, Somayyeh Fahiminiya, Spyros Oikonomopoulos, +1
Methods in Molecular Biology (16 May, 2018)
Transcript Profiling Using Long-Read Sequencing Technologies
by Anthony Bayega, Yu Chang Wang, Spyros Oikonomopoulos, +3
Methods in Molecular Biology (16 May, 2018)