Biology subjects

Pryszcz, L. P.

Publications and source records attributed to Pryszcz, L. P..

6 recordsLinked to original sources

bioRxiv2026

AMaNITA: an end-to-end workflow for native tRNA nanopore sequencing data analysis

Transfer RNA (tRNA) molecules serve as essential adapters during protein translation. While direct RNA sequencing (DRS) via Oxford Nanopore Technologies has emerged as a powerful platform for systematic tRNAome profiling, we currently lack a simple and robust statistical framework for nanopore tRNA data analyses. Here, we address this gap by developing AMaNITA (Abundance, Modifications, and Nanopore Intensity Toolbox Application), an end-to-end bioinformatic workflow that enables simplified, robust, and scalable analyses of nanopore native tRNA sequencing datasets. AMaNITA streamlines the entire analytical trajectory: from upstream processing (basecalling, mapping, filtering, batch effect correction) to downstream assessment of differential tRNA abundance and modification stoichiometry. The workflow generates an interactive HTML report for data exploration and analysis, allowing the user to download the source data files and resulting plots. AMaNITA can be executed using Singularity from the command line, without requiring installation of dependencies.

bioinformatics↗

bioRxiv2024

SeqTagger, a rapid and accurate tool to demultiplex direct RNA nanopore sequencing datasets

Nanopore direct RNA sequencing (DRS) enables direct measurement of RNA molecules, including their native RNA modifications, without prior conversion to cDNA. However, commercial methods for molecular barcoding of multiple DRS samples are lacking, and community-driven efforts, such as DeePlexiCon, are not compatible with newer RNA chemistry flowcells and the latest-generation GPU cards. To overcome these limitations, we introduce SeqTagger, a rapid and robust method that can demultiplex direct RNA sequencing datasets with 99% precision and 95% recall. We demonstrate the applicability of SeqTagger in both RNA002/R9.4 and RNA004/RNA chemistries and show its robust performance both for long and short RNA libraries, including custom libraries that do not contain standard poly-(A) tails, such as Nano-tRNAseq libraries. Finally, we demonstrate that increasing the multiplexing up to 96 barcodes yields highly accurate demultiplexing models. SeqTagger can be executed in a standalone manner or through the MasterOfPores NextFlow workflow. The availability of an efficient and simple multiplexing strategy improves the cost-effectiveness of this technology and facilitates the analysis of low-input biological samples.

genomics↗

bioRxiv2021

ModPhred: an integrative toolkit for the analysis and storage of nanopore sequencing DNA and RNA modification data

MotivationDNA and RNA modifications can now be identified using Nanopore sequencing. However, we currently lack a flexible software to efficiently encode, store, analyze and visualize DNA and RNA modification data. ResultsHere we present ModPhred, a versatile toolkit that facilitates DNA and RNA modification analysis from nanopore sequencing reads in a user-friendly manner. ModPhred integrates probabilistic DNA and RNA modification information within the FASTQ and BAM file formats, can be used to encode multiple types of modifications simultaneously, and its output can be easily coupled to genomic track viewers, facilitating the visualization and analysis of DNA and RNA modification information in individual reads in a simple and computationally efficient manner. Availability and ImplementationModPhred is available at https://github.com/novoalab/modPhred, is implemented in Python3, and is released under an MIT license. Supplementary DataSupplementary Data are available at Bioinformatics online.

bioinformatics↗

bioRxiv2019

Parallel and scalable workflow for the analysis of Oxford Nanopore direct RNA sequencing datasets

The direct RNA sequencing platform offered by Oxford Nanopore Technologies allows for direct measurement of RNA molecules without the need of conversion to complementary DNA, fragmentation or amplification. As such, it is virtually capable of detecting any given RNA modification present in the molecule that is being sequenced, as well as provide polyA tail length estimations at the level of individual RNA molecules. Although this technology has been publicly available since 2017, the complexity of the raw Nanopore data, together with the lack of systematic and reproducible pipelines, have greatly hindered the access of this technology to the general user. Here we address this problem by providing a fully benchmarked workflow for the analysis of direct RNA sequencing reads, termed MasterOfPores. The pipeline converts raw current intensities into multiple types of processed data, providing metrics of the quality of the run, quality-filtering, base-calling and mapping. The output of the pipeline can in turn be used to compute per-gene counts, RNA modifications, and prediction of polyA tail length and RNA isoforms. The software is written using the NextFlow framework for parallelization and portability, and relies on Linux containers such as Docker and Singularity for achieving better reproducibility. The MasterOfPores workflow can be executed on any Unix-compatible OS on a computer, cluster or cloud without the need of installing any additional software or dependencies, and is freely available in Github (https://github.com/biocorecrg/master_of_pores). This workflow will significantly simplify the analysis of nanopore direct RNA sequencing data by non-bioinformatics experts, thus boosting the understanding of the (epi)transcriptome with single molecule resolution.

bioinformatics↗

bioRxiv2017

nextPARS: Parallel probing of RNA structures in Illumina

RNA molecules play important roles in virtually every cellular process. These functions are often mediated through the adoption of specific structures that enable RNAs to interact with other molecules. Thus, determining the secondary structures of RNAs is central to understanding their function and evolution. In recent years several sequencing-based approaches have been developed that allow probing structural features of thousands of RNA molecules present in a sample. Here, we describe nextPARS, a novel Illumina-based implementation of in-vitro parallel probing of RNA structures. Our approach achieves comparable accuracy to previous implementations, while enabling higher throughput and sample multiplexing.

genomics↗

bioRxiv2017

Distinct roles of the polarity factors Boi1 and Boi2 in the control of exocytosis and abscission in budding yeast

Boi1 and Boi2 (Boi1/2) are budding yeast plasma membrane proteins that function in polarized growth, and in cytokinesis inhibition in response to chromosome bridges via the NoCut abscission checkpoint. How Boi1/2 act in these two distinct processes is not understood. We demonstrate that Boi1/2 are required for a late step in the fusion of secretory vesicles with the plasma membrane of the growing bud. Cells lacking Boi1/2 accumulate secretory vesicles and are defective in bud growth. In contrast, Boi2 is specifically required for abscission inhibition in cells with chromatin bridges. The SH3 domain of Boi2, which is dispensable for bud growth and targets Boi2 to the site of abscission, is essential for abscission inhibition. Gain of function of the exocyst, a conserved protein complex involved in tethering of exocytic vesicles to the plasma membrane, rescued secretion and bud growth defects in boi mutant cells, and abrogated NoCut checkpoint function. Thus, Boi2 functions redundantly with Boi1 to promote the fusion of secretory vesicles with the plasma membrane at sites of polarized growth, and acts as an abscission inhibitor during cytokinesis in response to chromatin bridges.

cell biology↗