Complete Transcriptome RNA-Seq

David F.B. Miller; Pearlly Yan; Fang Fang; Aaron Buechlein; Karl Kroll; David Frankhouser; Cameron Stump; Paige Stump; James B. Ford; Haixu Tang; Scott Michaels; Daniela Matei; Tim H. Huang; Jeremy Chien; Yunlong Liu; Douglas B. Rusch; Kenneth P. Nephew

doi:10.1007/978-1-4939-6539-7_10

Complete Transcriptome RNA-Seq

David F.B. Miller, Pearlly Yan, Fang Fang, Aaron Buechlein, Karl Kroll, David Frankhouser, Cameron Stump, Paige Stump, James B. Ford, Haixu Tang, Scott Michaels, Daniela Matei, Tim H. Huang, Jeremy Chien, Yunlong Liu, Douglas B. Rusch, Kenneth P. Nephew

Department of Molecular Medicine

Research output: Chapter in Book/Report/Conference proceeding › Chapter

3 Scopus citations

Abstract

RNA-Seq is the leading technology for analyzing gene expression on a global scale across a broad spectrum of sample types. However, due to chemical modifi cations by fi xation or degradation due to collection methods, samples often contain an abundance of RNA that is no longer intact, and the capability of current RNA-Seq protocols to accurately quantify such samples is often limited. We have developed an RNASeq protocol to address these key issues as well as quantify gene expression from the whole transcriptome. Furthermore, for compatibility with improved sequencing platforms, we use restructured adapter sequences to generate libraries for Illumina HiSeq, MiSeq, and NextSeq platforms. Our protocol utilizes duplexspecifi c nuclease (DSN) to remove abundant ribosomal RNA sequences while retaining other types of RNA for superior transcriptome profi ling from low quantity input. We employ the Illumina sequencing platform, but this method is described in suffi cient detail to adapt to other platforms.

Original language	English (US)
Title of host publication	Methods in Molecular Biology
Publisher	Humana Press
Pages	141-162
Number of pages	22
DOIs	https://doi.org/10.1007/978-1-4939-6539-7_10
State	Published - 2017

Publication series

Name	Methods in Molecular Biology
Volume	1513
ISSN (Print)	1064-3745

Keywords

Duplex-specifi c Nuclease
Gene Expression
RNA-Seq
Sequencing
Transcriptome

ASJC Scopus subject areas

Genetics
Molecular Biology

Access to Document

10.1007/978-1-4939-6539-7_10

Cite this

Miller, D. F. B., Yan, P., Fang, F., Buechlein, A., Kroll, K., Frankhouser, D., Stump, C., Stump, P., Ford, J. B., Tang, H., Michaels, S., Matei, D., Huang, T. H., Chien, J., Liu, Y., Rusch, D. B., & Nephew, K. P. (2017). Complete Transcriptome RNA-Seq. In Methods in Molecular Biology (pp. 141-162). (Methods in Molecular Biology; Vol. 1513). Humana Press. https://doi.org/10.1007/978-1-4939-6539-7_10

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abstract = "RNA-Seq is the leading technology for analyzing gene expression on a global scale across a broad spectrum of sample types. However, due to chemical modifi cations by fi xation or degradation due to collection methods, samples often contain an abundance of RNA that is no longer intact, and the capability of current RNA-Seq protocols to accurately quantify such samples is often limited. We have developed an RNASeq protocol to address these key issues as well as quantify gene expression from the whole transcriptome. Furthermore, for compatibility with improved sequencing platforms, we use restructured adapter sequences to generate libraries for Illumina HiSeq, MiSeq, and NextSeq platforms. Our protocol utilizes duplexspecifi c nuclease (DSN) to remove abundant ribosomal RNA sequences while retaining other types of RNA for superior transcriptome profi ling from low quantity input. We employ the Illumina sequencing platform, but this method is described in suffi cient detail to adapt to other platforms.",

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author = "Miller, {David F.B.} and Pearlly Yan and Fang Fang and Aaron Buechlein and Karl Kroll and David Frankhouser and Cameron Stump and Paige Stump and Ford, {James B.} and Haixu Tang and Scott Michaels and Daniela Matei and Huang, {Tim H.} and Jeremy Chien and Yunlong Liu and Rusch, {Douglas B.} and Nephew, {Kenneth P.}",

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doi = "10.1007/978-1-4939-6539-7_10",

language = "English (US)",

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AU - Yan, Pearlly

AU - Fang, Fang

AU - Buechlein, Aaron

AU - Kroll, Karl

AU - Frankhouser, David

AU - Stump, Cameron

AU - Stump, Paige

AU - Ford, James B.

AU - Tang, Haixu

AU - Michaels, Scott

AU - Matei, Daniela

AU - Huang, Tim H.

AU - Chien, Jeremy

AU - Liu, Yunlong

AU - Rusch, Douglas B.

AU - Nephew, Kenneth P.

PY - 2017

Y1 - 2017

N2 - RNA-Seq is the leading technology for analyzing gene expression on a global scale across a broad spectrum of sample types. However, due to chemical modifi cations by fi xation or degradation due to collection methods, samples often contain an abundance of RNA that is no longer intact, and the capability of current RNA-Seq protocols to accurately quantify such samples is often limited. We have developed an RNASeq protocol to address these key issues as well as quantify gene expression from the whole transcriptome. Furthermore, for compatibility with improved sequencing platforms, we use restructured adapter sequences to generate libraries for Illumina HiSeq, MiSeq, and NextSeq platforms. Our protocol utilizes duplexspecifi c nuclease (DSN) to remove abundant ribosomal RNA sequences while retaining other types of RNA for superior transcriptome profi ling from low quantity input. We employ the Illumina sequencing platform, but this method is described in suffi cient detail to adapt to other platforms.

AB - RNA-Seq is the leading technology for analyzing gene expression on a global scale across a broad spectrum of sample types. However, due to chemical modifi cations by fi xation or degradation due to collection methods, samples often contain an abundance of RNA that is no longer intact, and the capability of current RNA-Seq protocols to accurately quantify such samples is often limited. We have developed an RNASeq protocol to address these key issues as well as quantify gene expression from the whole transcriptome. Furthermore, for compatibility with improved sequencing platforms, we use restructured adapter sequences to generate libraries for Illumina HiSeq, MiSeq, and NextSeq platforms. Our protocol utilizes duplexspecifi c nuclease (DSN) to remove abundant ribosomal RNA sequences while retaining other types of RNA for superior transcriptome profi ling from low quantity input. We employ the Illumina sequencing platform, but this method is described in suffi cient detail to adapt to other platforms.

KW - Duplex-specifi c Nuclease

KW - Gene Expression

KW - RNA-Seq

KW - Sequencing

KW - Transcriptome

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PB - Humana Press

ER -

Complete Transcriptome RNA-Seq

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ASJC Scopus subject areas

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