Small-scale sequencing enables quality assessment of Ribo-Seq data: an example from Arabidopsis cell culture

Translation is a tightly regulated process, controlling the rate of protein synthesis in cells. Ribosome sequencing (Ribo-Seq) is a recently developed tool for studying actively translated mRNA and can thus directly address translational regulation. Ribo-Seq libraries need to be sequenced to a great...

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Bibliographic Details
Published in:Plant methods 2021-08, Vol.17 (1), p.92-92, Article 92
Main Authors: Mahboubi, Amir, Delhomme, Nicolas, Häggström, Sara, Hanson, Johannes
Format: Article
Language:English
Subjects:
Arabidopsis
Bioinformatics (Computational Biology)
Bioinformatik (beräkningsbiologi)
Cell culture
Contamination
Data analysis
Decision making
DNA sequencing
Evaluation
Evaluation of sequencing library quality
Gene expression
Genetic aspects
Genomes
Libraries
Methodology
Methods
mRNA
Nucleic acids
Nucleotide sequencing
Optimization
Periodicity
Physiological aspects
Pipelines
Plant tissue culture
Protein biosynthesis
Protein synthesis
Proteins
Qualitative analysis
Quality assessment
Quality control
Ribo-Seq
Ribosomal profiling
ribosomes
rRNA
Translation
Translational profiling
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Summary:Translation is a tightly regulated process, controlling the rate of protein synthesis in cells. Ribosome sequencing (Ribo-Seq) is a recently developed tool for studying actively translated mRNA and can thus directly address translational regulation. Ribo-Seq libraries need to be sequenced to a great depth due to high contamination by rRNA and other contaminating nucleic acid fragments. Deep sequencing is expensive, and it generates large volumes of data, making data analysis complicated and time consuming. Here we developed a platform for Ribo-Seq library construction and data analysis to enable rapid quality assessment of Ribo-Seq libraries with the help of a small-scale sequencer. Our data show that several qualitative features of a Ribo-Seq library, such as read length distribution, P-site distribution, reading frame and triplet periodicity, can be effectively evaluated using only the data generated by a benchtop sequencer with a very limited number of reads. Our pipeline enables rapid evaluation of Ribo-Seq libraries, opening up possibilities for optimization of Ribo-Seq library construction from difficult samples, and leading to better decision making prior to more costly deep sequencing.
ISSN:1746-4811
1746-4811
DOI:10.1186/s13007-021-00791-w