CoolMPS for robust sequencing of single-nuclear RNAs captured by droplet-based method.
Nucleic Acids Res
; 49(2): e11, 2021 01 25.
Article
em En
| MEDLINE
| ID: mdl-33264392
ABSTRACT
Massively-parallel single-cell and single-nucleus RNA sequencing (scRNA-seq, snRNA-seq) requires extensive sequencing to achieve proper per-cell coverage, making sequencing resources and availability of sequencers critical factors for conducting deep transcriptional profiling. CoolMPS is a novel sequencing-by-synthesis approach that relies on nucleotide labeling by re-usable antibodies, but whether it is applicable to snRNA-seq has not been tested. Here, we use a low-cost and off-the-shelf protocol to chemically convert libraries generated with the widely-used Chromium 10X technology to be sequenceable with CoolMPS technology. To assess the quality and performance of converted libraries sequenced with CoolMPS, we generated a snRNA-seq dataset from the hippocampus of young and old mice. Native libraries were sequenced on an Illumina Novaseq and libraries that were converted to be compatible with CoolMPS were sequenced on a DNBSEQ-400RS. CoolMPS-derived data faithfully replicated key characteristics of the native library dataset, including correct estimation of ambient RNA-contamination, detection of captured cells, cell clustering results, spatial marker gene expression, inter- and intra-replicate differences and gene expression changes during aging. In conclusion, our results show that CoolMPS provides a viable alternative to standard sequencing of RNA from droplet-based libraries.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
RNA Nuclear Pequeno
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Análise de Sequência de RNA
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Análise de Célula Única
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Sequenciamento de Nucleotídeos em Larga Escala
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Encapsulamento de Células
Limite:
Animals
Idioma:
En
Ano de publicação:
2021
Tipo de documento:
Article