LiBiNorm: an htseq-count analogue with improved normalisation of Smart-seq2 data and library preparation diagnostics

Protocols for preparing RNA sequencing (RNA-seq) libraries, most prominently “Smart-seq” variations, introduce global biases that can have a significant impact on the quantification of gene expression levels. This global bias can lead to drastic over- or under-representation of RNA in non-linear len...

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Vydáno v:	PeerJ (San Francisco, CA) Ročník 7; s. e6222
Hlavní autoři:	Dyer, Nigel P., Shahrezaei, Vahid, Hebenstreit, Daniel
Médium:	Journal Article
Jazyk:	angličtina
Vydáno:	United States PeerJ. Ltd 04.02.2019 PeerJ, Inc PeerJ Inc
Témata:	Bias Biochemistry Bioinformatics Biology Computational Biology Computational Science Computer programs Data processing Deoxyribonucleic acid DNA Enzymes Gene expression Genes Genetic research Genomes Genomics Global bias Methods Normalization Ribonucleic acid RNA RNA sequencing RNA-seq Scientific software Smart-seq2 Software Technology application Visualization (Computer) United Kingdom > UK Normalization Smart-seq2 RNA-seq Gene expression Global bias
ISSN:	2167-8359, 2167-8359
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Abstract	Protocols for preparing RNA sequencing (RNA-seq) libraries, most prominently “Smart-seq” variations, introduce global biases that can have a significant impact on the quantification of gene expression levels. This global bias can lead to drastic over- or under-representation of RNA in non-linear length-dependent fashion due to enzymatic reactions during cDNA production. It is currently not corrected by any RNA-seq software, which mostly focus on local bias in coverage along RNAs. This paper describes LiBiNorm, a simple command line program that mimics the popular htseq-count software and allows diagnostics, quantification, and global bias removal. LiBiNorm outputs gene expression data that has been normalized to correct for global bias introduced by the Smart-seq2 protocol. In addition, it produces data and several plots that allow insights into the experimental history underlying library preparation. The LiBiNorm package includes an R script that allows visualization of the main results. LiBiNorm is the first software application to correct for the global bias that is introduced by the Smart-seq2 protocol. It is freely downloadable at http://www2.warwick.ac.uk/fac/sci/lifesci/research/libinorm .
AbstractList	Protocols for preparing RNA sequencing (RNA-seq) libraries, most prominently “Smart-seq” variations, introduce global biases that can have a significant impact on the quantification of gene expression levels. This global bias can lead to drastic over- or under-representation of RNA in non-linear length-dependent fashion due to enzymatic reactions during cDNA production. It is currently not corrected by any RNA-seq software, which mostly focus on local bias in coverage along RNAs. This paper describes LiBiNorm, a simple command line program that mimics the popular htseq-count software and allows diagnostics, quantification, and global bias removal. LiBiNorm outputs gene expression data that has been normalized to correct for global bias introduced by the Smart-seq2 protocol. In addition, it produces data and several plots that allow insights into the experimental history underlying library preparation. The LiBiNorm package includes an R script that allows visualization of the main results. LiBiNorm is the first software application to correct for the global bias that is introduced by the Smart-seq2 protocol. It is freely downloadable at http://www2.warwick.ac.uk/fac/sci/lifesci/research/libinorm. Protocols for preparing RNA sequencing (RNA-seq) libraries, most prominently "Smart-seq" variations, introduce global biases that can have a significant impact on the quantification of gene expression levels. This global bias can lead to drastic over- or under-representation of RNA in non-linear length-dependent fashion due to enzymatic reactions during cDNA production. It is currently not corrected by any RNA-seq software, which mostly focus on local bias in coverage along RNAs. This paper describes LiBiNorm, a simple command line program that mimics the popular htseq-count software and allows diagnostics, quantification, and global bias removal. LiBiNorm outputs gene expression data that has been normalized to correct for global bias introduced by the Smart-seq2 protocol. In addition, it produces data and several plots that allow insights into the experimental history underlying library preparation. The LiBiNorm package includes an R script that allows visualization of the main results. LiBiNorm is the first software application to correct for the global bias that is introduced by the Smart-seq2 protocol. It is freely downloadable at http://www2.warwick.ac.uk/fac/sci/lifesci/research/libinorm.Protocols for preparing RNA sequencing (RNA-seq) libraries, most prominently "Smart-seq" variations, introduce global biases that can have a significant impact on the quantification of gene expression levels. This global bias can lead to drastic over- or under-representation of RNA in non-linear length-dependent fashion due to enzymatic reactions during cDNA production. It is currently not corrected by any RNA-seq software, which mostly focus on local bias in coverage along RNAs. This paper describes LiBiNorm, a simple command line program that mimics the popular htseq-count software and allows diagnostics, quantification, and global bias removal. LiBiNorm outputs gene expression data that has been normalized to correct for global bias introduced by the Smart-seq2 protocol. In addition, it produces data and several plots that allow insights into the experimental history underlying library preparation. The LiBiNorm package includes an R script that allows visualization of the main results. LiBiNorm is the first software application to correct for the global bias that is introduced by the Smart-seq2 protocol. It is freely downloadable at http://www2.warwick.ac.uk/fac/sci/lifesci/research/libinorm. Protocols for preparing RNA sequencing (RNA-seq) libraries, most prominently "Smart-seq" variations, introduce global biases that can have a significant impact on the quantification of gene expression levels. This global bias can lead to drastic over- or under-representation of RNA in non-linear length-dependent fashion due to enzymatic reactions during cDNA production. It is currently not corrected by any RNA-seq software, which mostly focus on local bias in coverage along RNAs. This paper describes LiBiNorm, a simple command line program that mimics the popular htseq-count software and allows diagnostics, quantification, and global bias removal. LiBiNorm outputs gene expression data that has been normalized to correct for global bias introduced by the Smart-seq2 protocol. In addition, it produces data and several plots that allow insights into the experimental history underlying library preparation. The LiBiNorm package includes an R script that allows visualization of the main results. LiBiNorm is the first software application to correct for the global bias that is introduced by the Smart-seq2 protocol. It is freely downloadable at Protocols for preparing RNA sequencing (RNA-seq) libraries, most prominently “Smart-seq” variations, introduce global biases that can have a significant impact on the quantification of gene expression levels. This global bias can lead to drastic over- or under-representation of RNA in non-linear length-dependent fashion due to enzymatic reactions during cDNA production. It is currently not corrected by any RNA-seq software, which mostly focus on local bias in coverage along RNAs. This paper describes LiBiNorm, a simple command line program that mimics the popular htseq-count software and allows diagnostics, quantification, and global bias removal. LiBiNorm outputs gene expression data that has been normalized to correct for global bias introduced by the Smart-seq2 protocol. In addition, it produces data and several plots that allow insights into the experimental history underlying library preparation. The LiBiNorm package includes an R script that allows visualization of the main results. LiBiNorm is the first software application to correct for the global bias that is introduced by the Smart-seq2 protocol. It is freely downloadable at http://www2.warwick.ac.uk/fac/sci/lifesci/research/libinorm .
ArticleNumber	e6222
Audience	Academic
Author	Hebenstreit, Daniel Dyer, Nigel P. Shahrezaei, Vahid
Author_xml	– sequence: 1 givenname: Nigel P. orcidid: 0000-0001-6158-0510 surname: Dyer fullname: Dyer, Nigel P. organization: School of Life Sciences, University of Warwick, Coventry, UK – sequence: 2 givenname: Vahid surname: Shahrezaei fullname: Shahrezaei, Vahid organization: Department of Mathematics, Imperial College London, London, UK – sequence: 3 givenname: Daniel surname: Hebenstreit fullname: Hebenstreit, Daniel organization: School of Life Sciences, University of Warwick, Coventry, UK
BackLink	https://www.ncbi.nlm.nih.gov/pubmed/30740268$$D View this record in MEDLINE/PubMed
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Cites_doi	10.1038/nprot.2016.095 10.1038/nmeth.1557 10.1186/gb-2013-14-4-r36 10.1016/j.celrep.2012.08.003 10.1038/nprot.2014.006 10.1186/s13059-014-0550-8 10.1101/035758 10.7717/peerj.869 10.1038/nmeth.3317 10.12688/f1000research.11290.1 10.1093/bioinformatics/btp692 10.1038/nmeth0205-151 10.1038/nmeth.4197 10.1201/b11812 10.1093/comjnl/7.4.308 10.1093/bioinformatics/bts635 10.1038/nbt.2282 10.1371/journal.pcbi.1005515 10.1038/nbt.1621 10.1093/bioinformatics/btp352 10.1016/j.cels.2016.10.012 10.1186/gb-2011-12-3-r22 10.1186/1471-2105-15-s1-s7 10.1186/gb-2013-14-4-r31 10.1093/bioinformatics/btu638 10.1038/nmeth.2772
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Snippet	Protocols for preparing RNA sequencing (RNA-seq) libraries, most prominently “Smart-seq” variations, introduce global biases that can have a significant impact... Protocols for preparing RNA sequencing (RNA-seq) libraries, most prominently "Smart-seq" variations, introduce global biases that can have a significant impact...
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SubjectTerms	Bias Biochemistry Bioinformatics Biology Computational Biology Computational Science Computer programs Data processing Deoxyribonucleic acid DNA Enzymes Gene expression Genes Genetic research Genomes Genomics Global bias Methods Normalization Ribonucleic acid RNA RNA sequencing RNA-seq Scientific software Smart-seq2 Software Technology application Visualization (Computer)
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Title	LiBiNorm: an htseq-count analogue with improved normalisation of Smart-seq2 data and library preparation diagnostics
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