Abstract
Single-cell DNA sequencing methods are challenged by poor physical coverage, high technical error rates and low throughput. To address these issues, we developed a single-cell DNA sequencing protocol that combines flow-sorting of single nuclei, time-limited multiple-displacement amplification (MDA), low-input library preparation, DNA barcoding, targeted capture and next-generation sequencing (NGS). This approach represents a major improvement over our previous single nucleus sequencing (SNS) Nature Protocols paper in terms of generating higher-coverage data (>90%), thereby enabling the detection of genome-wide variants in single mammalian cells at base-pair resolution. Furthermore, by pooling 48-96 single-cell libraries together for targeted capture, this approach can be used to sequence many single-cell libraries in parallel in a single reaction. This protocol greatly reduces the cost of single-cell DNA sequencing, and it can be completed in 5-6 d by advanced users. This single-cell DNA sequencing protocol has broad applications for studying rare cells and complex populations in diverse fields of biological research and medicine.
Original language | English (US) |
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Pages (from-to) | 214-235 |
Number of pages | 22 |
Journal | Nature protocols |
Volume | 11 |
Issue number | 2 |
DOIs | |
State | Published - Feb 1 2016 |
ASJC Scopus subject areas
- General Biochemistry, Genetics and Molecular Biology
MD Anderson CCSG core facilities
- Advanced Technology Genomics Core
- Bioinformatics Shared Resource
- Flow Cytometry and Cellular Imaging Facility