Single-cell RNA-seq analysis of mouse preimplantation embryos by third-generation sequencing.
Single-cell RNA-seq analysis of mouse preimplantation embryos by third-generation sequencing.
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DOI：
10.1371/journal.pbio.3001017
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发表时间：
2020-12
期刊：
影响因子：
9.8
通讯作者：
Tang F
中科院分区：
文献类型：
--
作者：
Fan X;Tang D;Liao Y;Li P;Zhang Y;Wang M;Liang F;Wang X;Gao Y;Wen L;Wang D;Wang Y;Tang F
基于下一代测序（NGS）平台的单细胞RNA测序（scRNA-seq）技术的发展极大地改变了生物学研究，但由于其读长较短，仍然存在许多问题无法解决。我们开发了一种基于第三代测序（TGS）平台的新型scRNA-seq技术（通过Nanopore平台对全长RNA进行单细胞扩增和测序，SCAN-seq）。 SCAN-seq 表现出与基于 NGS 平台的 scRNA-seq 方法相当的高灵敏度和准确性。此外，我们在小鼠胚胎干细胞（mESC）中通过逆转录PCR（RT-PCR）耦合桑格测序捕获了数千个不同类型的未注释转录本，并具有高验证率。然后，我们使用 SCAN-seq 分析小鼠植入前胚胎。我们可以清楚地区分不同发育阶段的细胞，共鉴定出来自 9,338 个基因的 27,250 个未注释转录本，其中许多转录本表现出发育阶段特异性的表达模式。最后，我们表明 SCAN-seq 在确定单个细胞内等位基因特异性基因表达模式方面表现出很高的准确性。 SCAN-seq在单细胞转录组分析领域取得重大突破。本研究描述了一种名为SCAN-seq的新型单细胞RNA-seq技术，该技术基于第三代Nanopore测序平台可以捕获单细胞中的全长转录本，并在小鼠植入前胚胎上展示了其性能。
The development of next generation sequencing (NGS) platform-based single-cell RNA sequencing (scRNA-seq) techniques has tremendously changed biological researches, while there are still many questions that cannot be addressed by them due to their short read lengths. We developed a novel scRNA-seq technology based on third-generation sequencing (TGS) platform (single-cell amplification and sequencing of full-length RNAs by Nanopore platform, SCAN-seq). SCAN-seq exhibited high sensitivity and accuracy comparable to NGS platform-based scRNA-seq methods. Moreover, we captured thousands of unannotated transcripts of diverse types, with high verification rate by reverse transcription PCR (RT-PCR)–coupled Sanger sequencing in mouse embryonic stem cells (mESCs). Then, we used SCAN-seq to analyze the mouse preimplantation embryos. We could clearly distinguish cells at different developmental stages, and a total of 27,250 unannotated transcripts from 9,338 genes were identified, with many of which showed developmental stage-specific expression patterns. Finally, we showed that SCAN-seq exhibited high accuracy on determining allele-specific gene expression patterns within an individual cell. SCAN-seq makes a major breakthrough for single-cell transcriptome analysis field. This study describes a novel single-cell RNA-seq technology called SCAN-seq which can capture the full-length transcripts in single cells based on the third-generation Nanopore sequencing platform, and demonstrates its performance on mouse preimplantation embryos.
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影响因子：
--
作者：
Codina-Fauteux, Valerie-Anne;Beaudoin, Melissa;Lettre, Guillaume
通讯作者：
Lettre, Guillaume
影响因子：
8.8
作者：
Cui, Yueli;Zheng, Yuxuan;Tang, Fuchou
通讯作者：
Tang, Fuchou
影响因子：
16.6
作者：
Byrne A;Beaudin AE;Olsen HE;Jain M;Cole C;Palmer T;DuBois RM;Forsberg EC;Akeson M;Vollmers C
通讯作者：
Vollmers C
影响因子：
7
作者：
Tardaguila M;de la Fuente L;Marti C;Pereira C;Pardo-Palacios FJ;Del Risco H;Ferrell M;Mellado M;Macchietto M;Verheggen K;Edelmann M;Ezkurdia I;Vazquez J;Tress M;Mortazavi A;Martens L;Rodriguez-Navarro S;Moreno-Manzano V;Conesa A
通讯作者：
Conesa A
DOI：
10.1007/978-1-4939-9240-9_6
发表时间：
2019-01-01
期刊：
SINGLE CELL METHODS: SEQUENCING AND PROTEOMICS
影响因子：
--
作者：
Bageritz, Josephine;Raddi, Gianmarco
通讯作者：
Raddi, Gianmarco