Abstract |
We show the use of 5'-Acrydite oligonucleotides to copolymerize single-cell DNA or RNA into balls of acrylamide gel (BAGs). Combining this step with split-and-pool techniques for creating barcodes yields a method with advantages in cost and scalability, depth of coverage, ease of operation, minimal cross-contamination, and efficient use of samples. We perform DNA copy number profiling on mixtures of cell lines, nuclei from frozen prostate tumors, and biopsy washes. As applied to RNA, the method has high capture efficiency of transcripts and sufficient consistency to clearly distinguish the expression patterns of cell lines and individual nuclei from neurons dissected from the mouse brain. By using varietal tags (UMIs) to achieve sequence error correction, we show extremely low levels of cross-contamination by tracking source-specific SNVs. The method is readily modifiable, and we will discuss its adaptability and diverse applications.
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Authors | Siran Li, Jude Kendall, Sarah Park, Zihua Wang, Joan Alexander, Andrea Moffitt, Nissim Ranade, Cassidy Danyko, Bruno Gegenhuber, Stephan Fischer, Brian D Robinson, Herbert Lepor, Jessica Tollkuhn, Jesse Gillis, Eric Brouzes, Alex Krasnitz, Dan Levy, Michael Wigler |
Journal | Genome research
(Genome Res)
Vol. 30
Issue 1
Pg. 49-61
(01 2020)
ISSN: 1549-5469 [Electronic] United States |
PMID | 31727682
(Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)
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Copyright | © 2020 Li et al.; Published by Cold Spring Harbor Laboratory Press. |
Chemical References |
- Nucleic Acids
- Acrylamide
- RNA
- DNA
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Topics |
- Acrylamide
(chemistry)
- DNA
- DNA Contamination
- DNA Copy Number Variations
- Gene Dosage
- Gene Expression Profiling
(methods, standards)
- Gene Library
- Humans
- Neoplasms
(genetics, metabolism, pathology)
- Nucleic Acids
(chemistry)
- Oligonucleotide Array Sequence Analysis
(methods, standards)
- Polymerization
- RNA
- Single-Cell Analysis
(methods, standards)
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