IsoQuant: a tool for accurate novel isoform discovery with long reads

Prjibelski, Andrey D.; Mikheenko, Alla; Joglekar, Anoushka; Smetanin, Alexander; Jarroux, Julien; Lapidus, Alla; Tilgner, Hagen U

doi:10.21203/rs.3.rs-1571850/v1

Cited by 10 publications

(9 citation statements)

References 17 publications

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“…To determine which HIV-1 transcripts were detected, we next mapped individual reads from the untreated sample to the HIV-1 transcriptome using IsoQuant 36 (See Material and Methods). Across all samples, over 80% of reads were unambiguously assigned to a single known isoform (Sup.…”

Section: Resultsmentioning

confidence: 99%

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Nano-DMS-MaP-seq allows isoform specific RNA structure determination

Smyth

Bohn

Gribling-Burrer

et al. 2022

Preprint

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Chemical probing experiments interrogate RNA structure by reacting with unpaired residues whose positions are typically read out by mutational profiling on short-read sequencing machines. Such experiments make rapid and genome-wide measurements of RNA structure, but short sequencing reads can rarely be mapped to specific transcript isoforms. Consequently, RNA structure information is acquired as a population average in common regions between transcripts. Here, we present nanopore dimethyl-sulfate mutational profiling (Nano-DMS-MaP-seq) for obtaining isoform resolved structural information of highly similar RNA molecules. We demonstrate the utility of Nano-DMS-MaP-seq by resolving the complex structural landscape of HIV-1 transcripts in infected cells. We show that unspliced and spliced transcripts have distinct structures in the common 5’UTR explaining the specific packaging of only unspliced RNAs into viral particles. Thus, Nano-DMS-MaP-seq is a straightforward method to resolve biologically important transcript-specific RNA structures that were previously hidden in short read ensemble analyses.

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Section: Resultsmentioning

confidence: 99%

“…Read to isoform mapping was performed using IsoQuant 36 , using a general feature le (GFF) format generated from previously published data 31 , but adjusted for PCR primer start and end sites. Only reads mapping uniquely to one isoform were sub-sorted for subsequent analyses.…”

Section: Basecalling and Isoform Detectionmentioning

confidence: 99%

Nano-DMS-MaP-seq allows isoform specific RNA structure determination

Smyth

Bohn

Gribling-Burrer

et al. 2022

Preprint

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“…We compared IsoTools transcriptome reconstruction with four other recently published tools (IsoQuant [22], TALON [14] , FLAIR [21] and Bambu [23]). Each tool was applied to the hepatocyte data set with its default filtering criteria, and IsoTools transcript reconstruction was run with varying filters.…”

Section: Lrts Transcriptome Reconstruction Is Largely Dependent On Fi...mentioning

confidence: 99%

“…TALON was developed as the ENCODE4 pipeline for long-read sequencing, and provides similar functionality to SQANTI3 [14].The Swan library [16], which is also part of the ENCODE4 pipeline, implements a statistical test based on a negative binomial model for differential expression analysis on isoform level using LRTS. Further tools for transcriptome reconstruction include Flair [21], IsoQuant [22] and Bambu [23].…”

Section: Introductionmentioning

confidence: 99%

IsoTools – a ﬂexible workﬂow for long-read transcriptome sequencing analysis

Lienhard

Beucken

Timmermann

et al. 2022

Preprint

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Long-read transcriptome sequencing (LRTS) holds the promise to reﬁne our understanding of alternative splicing, however, the nature of splicing is highly complex, and computational tools need to cope with this complexity by maximum ﬂexibility at diﬀerent steps of the work-ﬂow. For this purpose we developed IsoTools, a Python-based LRTS analysis package. IsoTools provides broad functionality for transcriptome reconstruction and quantiﬁcation of isoforms. Additionally, we implemented a graph-based method for the identiﬁcation of alternative splicing events and a statistical approach based on the beta-binomial distribution for the detection of diﬀerential events. To demonstrate our methods, we generated PacBio LRTS data of human hepatocytes treated with the HDAC inhibitor valproic acid. Contrasted with short read RNA-seq, this analysis shows that LRTS provides valuable additional insights for a better understanding of alternative splicing, in particular with respect to complex novel and diﬀerential splicing patterns. IsoTools is available at https://github.com/MatthiasLienhard/isotools.

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“…However many of these tools rely heavily on previously generated genome annotations. As part of the LRGASP consortium, several isoform identification tools, including StringTie [3,12], IsoQuant [13], IsoTools [14], Bambu [15], FLAIR [16], FLAMES [17], TALON [18], and our own Mandalorion were compared by a group of independent evaluators to assess performance [19]. Mandalorion performed very well compared to these other tools, likely driven by high Recall when identifying non-annotated isoforms.…”

Section: Introductionmentioning

confidence: 99%

Identifying and quantifying isoforms from accurate full-length transcriptome sequencing reads with Mandalorion

Volden

Schimke

Byrne

et al. 2022

Preprint

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The Mandalorion tool, which we have continuously developed over the last 5 years, identifies and quantifies high-confidence isoforms from accurate full-length transcriptome sequencing reads produced by methods like PacBio Iso-Seq and ONT-based R2C2. In this manuscript, we introduce and benchmark Mandalorion v4 which further improves upon the already strong performance of Mandalorion v3.6 used in the LRGASP consortium challenge. By processing real and simulated accurate full-length transcriptome sequencing data sets, we show three main features of Mandalorion: First, Mandalorion-based isoform identification has very high Precision and maintains high Recall even when used in the absence of any genome annotation. Second, isoform read counts as quantified by Mandalorion show high correlation with simulated read counts. Third, isoforms identified by Mandalorion closely reflect the full-length transcriptome sequencing data sets they are based on.

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IsoQuant: a tool for accurate novel isoform discovery with long reads

Cited by 10 publications

References 17 publications

Nano-DMS-MaP-seq allows isoform specific RNA structure determination

Nano-DMS-MaP-seq allows isoform specific RNA structure determination

IsoTools – a ﬂexible workﬂow for long-read transcriptome sequencing analysis

Identifying and quantifying isoforms from accurate full-length transcriptome sequencing reads with Mandalorion

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