BaRTv2: a Highly Resolved Barley Reference Transcriptome for Accurate Transcript-specific RNA-seq Quantification

Overview

Journal Plant J

Specialties Biology
Cell Biology
Molecular Biology

Date 2022 Jun 15

PMID 35704392

Authors

Max Coulter

Juan Carlos Entizne

Wenbin Guo

Micha Bayer

Ronja Wonneberger

Linda Milne

Miriam Schreiber

Allison Haaning

Gary J Muehlbauer

Nicola McCallum

John Fuller

Craig Simpson

Nils Stein

John W S Brown

Robbie Waugh

Runxuan Zhang

Affiliations

Soon will be listed here.

Abstract

Accurate characterisation of splice junctions (SJs) as well as transcription start and end sites in reference transcriptomes allows precise quantification of transcripts from RNA-seq data, and enables detailed investigations of transcriptional and post-transcriptional regulation. Using novel computational methods and a combination of PacBio Iso-seq and Illumina short-read sequences from 20 diverse tissues and conditions, we generated a comprehensive and highly resolved barley reference transcript dataset from the European 2-row spring barley cultivar Barke (BaRTv2.18). Stringent and thorough filtering was carried out to maintain the quality and accuracy of the SJs and transcript start and end sites. BaRTv2.18 shows increased transcript diversity and completeness compared with an earlier version, BaRTv1.0. The accuracy of transcript level quantification, SJs and transcript start and end sites have been validated extensively using parallel technologies and analysis, including high-resolution reverse transcriptase-polymerase chain reaction and 5'-RACE. BaRTv2.18 contains 39 434 genes and 148 260 transcripts, representing the most comprehensive and resolved reference transcriptome in barley to date. It provides an important and high-quality resource for advanced transcriptomic analyses, including both transcriptional and post-transcriptional regulation, with exceptional resolution and precision.

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References

Brown J, Calixto C, Zhang R . High-quality reference transcript datasets hold the key to transcript-specific RNA-sequencing analysis in plants. New Phytol. 2016; 213(2):525-530. DOI: 10.1111/nph.14208. View

Chen L, Shi X, Nian B, Duan S, Jiang B, Wang X . Alternative Splicing Regulation of Anthocyanin Biosynthesis in var. Unveiled by PacBio Iso-Seq. G3 (Bethesda). 2020; 10(8):2713-2723. PMC: 7407465. DOI: 10.1534/g3.120.401451. View

Simao F, Waterhouse R, Ioannidis P, Kriventseva E, Zdobnov E . BUSCO: assessing genome assembly and annotation completeness with single-copy orthologs. Bioinformatics. 2015; 31(19):3210-2. DOI: 10.1093/bioinformatics/btv351. View

Pertea M, Pertea G, Antonescu C, Chang T, Mendell J, Salzberg S . StringTie enables improved reconstruction of a transcriptome from RNA-seq reads. Nat Biotechnol. 2015; 33(3):290-5. PMC: 4643835. DOI: 10.1038/nbt.3122. View

Mascher M, Gundlach H, Himmelbach A, Beier S, Twardziok S, Wicker T . A chromosome conformation capture ordered sequence of the barley genome. Nature. 2017; 544(7651):427-433. DOI: 10.1038/nature22043. View

Abdel-Ghany S, Hamilton M, Jacobi J, Ngam P, Devitt N, Schilkey F . A survey of the sorghum transcriptome using single-molecule long reads. Nat Commun. 2016; 7:11706. PMC: 4931028. DOI: 10.1038/ncomms11706. View

Zhang Y, Gu L, Hou Y, Wang L, Deng X, Hang R . Integrative genome-wide analysis reveals HLP1, a novel RNA-binding protein, regulates plant flowering by targeting alternative polyadenylation. Cell Res. 2015; 25(7):864-76. PMC: 4493284. DOI: 10.1038/cr.2015.77. View

Marquez Y, Brown J, Simpson C, Barta A, Kalyna M . Transcriptome survey reveals increased complexity of the alternative splicing landscape in Arabidopsis. Genome Res. 2012; 22(6):1184-95. PMC: 3371709. DOI: 10.1101/gr.134106.111. View

Ren P, Meng Y, Li B, Ma X, Si E, Lai Y . Molecular Mechanisms of Acclimatization to Phosphorus Starvation and Recovery Underlying Full-Length Transcriptome Profiling in Barley ( L.). Front Plant Sci. 2018; 9:500. PMC: 5915550. DOI: 10.3389/fpls.2018.00500. View

10.

Wang M, Wang P, Liang F, Ye Z, Li J, Shen C . A global survey of alternative splicing in allopolyploid cotton: landscape, complexity and regulation. New Phytol. 2017; 217(1):163-178. DOI: 10.1111/nph.14762. View

11.

Schwacke R, Ponce-Soto G, Krause K, Bolger A, Arsova B, Hallab A . MapMan4: A Refined Protein Classification and Annotation Framework Applicable to Multi-Omics Data Analysis. Mol Plant. 2019; 12(6):879-892. DOI: 10.1016/j.molp.2019.01.003. View

12.

Kozak M . An analysis of 5'-noncoding sequences from 699 vertebrate messenger RNAs. Nucleic Acids Res. 1987; 15(20):8125-48. PMC: 306349. DOI: 10.1093/nar/15.20.8125. View

13.

Proudfoot N . Ending the message: poly(A) signals then and now. Genes Dev. 2011; 25(17):1770-82. PMC: 3175714. DOI: 10.1101/gad.17268411. View

14.

Au K, Underwood J, Lee L, Wong W . Improving PacBio long read accuracy by short read alignment. PLoS One. 2012; 7(10):e46679. PMC: 3464235. DOI: 10.1371/journal.pone.0046679. View

15.

Baralle F, Giudice J . Alternative splicing as a regulator of development and tissue identity. Nat Rev Mol Cell Biol. 2017; 18(7):437-451. PMC: 6839889. DOI: 10.1038/nrm.2017.27. View

16.

Salmela L, Walve R, Rivals E, Ukkonen E . Accurate self-correction of errors in long reads using de Bruijn graphs. Bioinformatics. 2016; 33(6):799-806. PMC: 5351550. DOI: 10.1093/bioinformatics/btw321. View

17.

Mao S, Pachter L, Tse D, Kannan S . RefShannon: A genome-guided transcriptome assembler using sparse flow decomposition. PLoS One. 2020; 15(6):e0232946. PMC: 7266320. DOI: 10.1371/journal.pone.0232946. View

18.

Zhang R, Calixto C, Marquez Y, Venhuizen P, Tzioutziou N, Guo W . A high quality Arabidopsis transcriptome for accurate transcript-level analysis of alternative splicing. Nucleic Acids Res. 2017; 45(9):5061-5073. PMC: 5435985. DOI: 10.1093/nar/gkx267. View

19.

Salmela L, Rivals E . LoRDEC: accurate and efficient long read error correction. Bioinformatics. 2014; 30(24):3506-14. PMC: 4253826. DOI: 10.1093/bioinformatics/btu538. View

20.

Bolger A, Lohse M, Usadel B . Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics. 2014; 30(15):2114-20. PMC: 4103590. DOI: 10.1093/bioinformatics/btu170. View