A novel strand-specific RNA-sequencing protocol using dU-adaptor-assembled Tn5
© The Author(s) 2022. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissionsoup.com.
| Publié dans: | Journal of experimental botany. - 1985. - 74(2023), 6 vom: 28. März, Seite 1806-1820 |
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| Auteur principal: | |
| Autres auteurs: | , , , , , , , |
| Format: | Article en ligne |
| Langue: | English |
| Publié: |
2023
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| Accès à la collection: | Journal of experimental botany |
| Sujets: | Journal Article Research Support, Non-U.S. Gov't Tn5 dU-Tn5 dUTP lncRNA soybean stranded RNA-seq DNA, Complementary Chromatin plus... |
| Résumé: | © The Author(s) 2022. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissionsoup.com. Strand-specific RNA-seq is a powerful tool for the discovery of novel transcripts, annotation of genomes, and profiling of gene expression levels. Tn5 transposase has been successfully applied in massive-scale sequencing projects; in particular, Tn5 adaptor modification is used in epigenetics, genomic structure, and chromatin visualization. We developed a novel dU-adaptor-assembled Tn5-mediated strand-specific RNA-sequencing protocol and compared this method with the leading dUTP method in terms of experimental procedure and multiple quality metrics of the generated libraries. The results showed that the dU-Tn5 method is easy to operate and generates a strand-specific RNA-seq library of comparable quality considering library complexity, strand-specificity, evenness, and continuity of annotated transcript coverage. We also evaluated the performance of the dU-Tn5 method in identifying nitrogen-responsive protein-coding genes and long non-coding RNAs in soybean roots. The results indicated that ~62-70% of differentially expressed genes detected from conventional libraries were also detected in dU-Tn5 libraries, indicating good agreement of our method with the current standard; moreover, their fold-changes were highly correlated (R>0.9). Thus, our method provides a promising 'do-it-yourself' stranded RNA-seq procedure for gene expression profiling |
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| Description: | Date Completed 30.03.2023 Date Revised 04.04.2023 published: Print Citation Status MEDLINE |
| ISSN: | 1460-2431 |
| DOI: | 10.1093/jxb/erac515 |