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Status |
Public on Jul 21, 2020 |
Title |
Massively parallel and time-resolved RNA sequencing in single cells with scNT-Seq |
Organisms |
Homo sapiens; Mus musculus |
Experiment type |
Expression profiling by high throughput sequencing
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Summary |
Single-cell RNA sequencing offers snapshots of whole transcriptomes but obscures the temporal RNA dynamics. Here we present single-cell metabolically labeled new RNA tagging sequencing (scNT-Seq), a method for massively parallel analysis of newly-transcribed and pre-existing mRNAs from the same cell. This droplet microfluidics-based method enables high-throughput chemical conversion on barcoded beads, efficiently marking newly-transcribed mRNAs with T-to-C substitutions. With scNT-Seq, we jointly profiled new and old transcriptomes in ~55,000 single cells. These data revealed time-resolved transcription factor activities and cell state trajectories at single-cell level in response to neuronal activation. We further determined rates of RNA biogenesis and decay to uncover RNA regulatory strategies during stepwise conversion between pluripotent and rare totipotent two-cell-embryo-like (2C-like) stem cell states. Finally, integrating scNT-Seq with genetic perturbation identifies DNA methylcytosine dioxygenases as an epigenetic barrier into 2C-like cell state. Time-resolved single-cell transcriptomic analysis thus opens new lines of inquiry regarding cell-type-specific RNA regulatory mechanisms.
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Overall design |
Single-cell metabolically labeled new RNA tagging sequencing
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Contributor(s) |
Qiu Q, Hu P, Qiu X, Govek K, Gonzalez-Camara P, Wu H |
Citation(s) |
32868927 |
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Submission date |
Dec 11, 2019 |
Last update date |
Sep 26, 2020 |
Contact name |
Peng Hu |
E-mail(s) |
penghu@upenn.edu
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Organization name |
University of Pennsylvanian
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Department |
Genetics
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Lab |
Hao Wu
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Street address |
415 Curie Blvd, 545 CRB
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City |
Philadelphia |
State/province |
PA |
ZIP/Postal code |
19104 |
Country |
USA |
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Platforms (3) |
GPL18573 |
Illumina NextSeq 500 (Homo sapiens) |
GPL19057 |
Illumina NextSeq 500 (Mus musculus) |
GPL19415 |
Illumina NextSeq 500 (Homo sapiens; Mus musculus) |
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Samples (38)
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Relations |
BioProject |
PRJNA594978 |
SRA |
SRP236894 |