Rizzetto, Simone and Eltahla, Auda A. and Lin, Peijie and Bull, Rowena and Lloyd, Andrew R and Ho, Joshua W K and Venturi, Vanessa and Luciani, Fabio (2017) Impact of sequencing depth and read length on single cell RNA sequencing data of T cells. Scientific Reports, 7 (1). ISSN 2045-2322 (OA)
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Abstract
Single cell RNA sequencing (scRNA-seq) provides great potential in measuring the gene expression profiles of heterogeneous cell populations. In immunology, scRNA-seq allowed the characterisation of transcript sequence diversity of functionally relevant T cell subsets, and the identification of the full length T cell receptor (TCRαβ), which defines the specificity against cognate antigens. Several factors, e.g. RNA library capture, cell quality, and sequencing output affect the quality of scRNA-seq data. We studied the effects of read length and sequencing depth on the quality of gene expression profiles, cell type identification, and TCRαβ reconstruction, utilising 1,305 single cells from 8 publically available scRNA-seq datasets, and simulation-based analyses. Gene expression was characterised by an increased number of unique genes identified with short read lengths (<50 bp), but these featured higher technical variability compared to profiles from longer reads. Successful TCRαβ reconstruction was achieved for 6 datasets (81% - 100%) with at least 0.25 millions (PE) reads of length >50 bp, while it failed for datasets with <30 bp reads. Sufficient read length and sequencing depth can control technical noise to enable accurate identification of TCRαβ and gene expression profiles from scRNA-seq data of T cells.
| Item Type: | Article |
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| Additional Information: | Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. |
| Subjects: | R Medicine > R Medicine (General) |
| Depositing User: | Repository Administrator |
| Date Deposited: | 09 Oct 2017 00:36 |
| Last Modified: | 09 Oct 2017 00:36 |
| URI: | https://eprints.victorchang.edu.au/id/eprint/653 |
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