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| Status |
Public on May 11, 2020 |
| Title |
A single-cell and single-nucleus RNA-Seq toolbox for fresh and frozen human tumors |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
Single cell genomics is essential to chart tumor ecosystems. While single cell RNA-Seq (scRNA-Seq) profiles RNA from cells dissociated from fresh tumors, single nucleus RNA-Seq (snRNA-Seq) is needed to profile frozen or hard-to-dissociate tumors. Each requires customization to different tissue and tumor types, posing a barrier to adoption. Here, we developed a systematic toolbox for profiling fresh and frozen clinical tumor samples using scRNA-Seq and snRNA-Seq, respectively. We analyzed 212,498 cells and nuclei from 39 samples across 23 specimens spanning eight tumor types of varying tissue and sample characteristics. We evaluated protocols by cell and nucleus quality, recovery rate, and cellular composition. scRNA-Seq and snRNA-Seq from matched samples recovered the same cell types, but at different proportions. Our work provides guidance for studies in a broad range of tumors, including criteria for testing and selecting methods from the toolbox for other tumors, thus paving the way for charting tumor atlases.
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| Overall design |
We developed a systematic toolbox for fresh and frozen tumor processing using single cell (sc) and single nucleus (sn) RNA-Seq, respectively (Fig. 1a). The toolbox contains our experimental workflow and methods, computational pipelines, and evaluation metrics. To generalize across tumor and sample types, we tested eight tumor types with different tissue characteristics (Fig. 1b), including comparisons of matched fresh and frozen preparations from the same tumor specimen. We tested varying tissue and sample characteristics including resection, biopsy, ascites, and orthotopic patient-derived xenograft. We included samples from non-small cell lung carcinoma (NSCLC), metastatic breast cancer (MBC), ovarian cancer, neuroblastoma, glioblastoma (GBM), pediatric high-grade glioma, chronic lymphocytic leukemia (CLL), pediatric sarcoma, and melanoma (Fig. 1b). In total, we analyzed 212,498 cells and nuclei across 23 tumors, from 22 patients spanning 39 sample preparations. Our work provides direct recommended protocols for multiple tumor types, decisions trees that allow researchers to choose the most suitable protocol for their research goals, and guidelines on how to customize protocols for new tumor and specimen types.
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*Raw sequencing data is is available in the controlled access repository DUOS (https://duos.broadinstitute.org/), under DUOS Dataset IDs DUOS-000111, DUOS-000112, DUOS-000113 and DUOS-000114*
**Submitter declares reads will be made available through dbGaP.**
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| Contributor(s) |
Slyper M, Porter CM, Ashenberg O, Waldman J, Drokhlyansky E, Wakiro I, Smillie C, Smith-Rosario G, Wu J, Dionne D, Vigneau S, Jané-Valbuena J, Napolitano S, Su M, Patel AG, Karlstrom A, Gritsch S, Nomura M, Waghray A, Gohil SH, Tsankov AM, Jerby-Arnon L, Cohen O, Klughammer J, Rosen Y, Gould J, Nguyen L, Hofree M, Li B, Wu CJ, Izar B, Haq R, Hodi FS, Yoon CH, Hata AN, Baker SJ, Suvà ML, Bueno R, Stover EH, Clay MR, Dyer MA, Collins NB, Wagle N, Johnson BE, Rotem A, Rozenblatt-Rosen O, Regev A |
| Citation(s) |
32405060 |
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Michal Slyper, Caroline B. M. Porter, Orr Ashenberg, Julia Waldman, Eugene Drokhlyansky, Isaac Wakiro, Christopher Smillie, Gabriela Smith-Rosario, Jingyi Wu, Danielle Dionne, Sébastien Vigneau, Judit Jané-Valbuena, Timothy L. Tickle, Sara Napolitano, Mei-Ju Su, Anand G. Patel, Asa Karlstrom, Simon Gritsch, Masashi Nomura, Avinash Waghray, Satyen H. Gohil, Alexander M. Tsankov, Livnat Jerby-Arnon, Ofir Cohen, Johanna Klughammer, Yanay Rosen, Joshua Gould, Lan Nguyen, Matan Hofree, Peter J. Tramontozzi, Bo Li, Catherine J. Wu, Benjamin Izar, Rizwan Haq, F. Stephen Hodi, Charles H. Yoon, Aaron N. Hata, Suzanne J. Baker, Mario L. Suvà, Raphael Bueno, Elizabeth H. Stover, Michael R. Clay, Michael A. Dyer, Natalie B. Collins, Ursula A. Matulonis, Nikhil Wagle, Bruce E. Johnson, Asaf Rotem, and Orit Rozenblatt-Rosen. A single-cell and single-nucleus RNA-Seq toolbox for fresh and frozen human tumors. Nat Med 2020;26:792-802. doi:10.1038/s41591-020-0844-1
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| Submission date |
Nov 21, 2019 |
| Last update date |
Aug 26, 2020 |
| Contact name |
Orr Ashenberg |
| Organization name |
Broad Institute
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| Department |
Klarman Cell Observatory
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| Lab |
Aviv Regev
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| Street address |
415 Main Street
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| City |
Cambridge |
| State/province |
MA |
| ZIP/Postal code |
02142 |
| Country |
USA |
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| Platforms (1) |
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| Samples (40)
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| Relations |
| BioProject |
PRJNA591014 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE140819_RAW.tar |
1.1 Gb |
(http)(custom) |
TAR (of CSV, H5) |
| Raw data not provided for this record |
| Processed data provided as supplementary file |
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