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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
|
| 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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| |
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| Overall design |
Single-cell metabolically labeled new RNA tagging sequencing
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| |
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| Contributor(s) |
Qiu Q, Hu P, Qiu X, Govek K, Gonzalez-Camara P, Wu H |
| Citation(s) |
32868927 |
| Submission date |
Dec 11, 2019 |
| Last update date |
Sep 26, 2020 |
| Contact name |
Peng Hu |
| E-mail(s) |
penghu@upenn.edu
|
| Organization name |
University of Pennsylvanian
|
| Department |
Genetics
|
| Lab |
Hao Wu
|
| Street address |
415 Curie Blvd, 545 CRB
|
| 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) |
|
| Samples (38)
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| Relations |
| BioProject |
PRJNA594978 |
| SRA |
SRP236894 |
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