title: Highly multiplexed targeted DNA sequencing from single nuclei.
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.
authors.affiliation: Department of Genetics, University of Texas MD Anderson Cancer Center, Houston, Texas, USA.|Graduate Program in Genes and Development, Graduate School of Biomedical Sciences, University of Texas Health Science Center at Houston, Houston, Texas, USA.
authors.forename: Marco L
authors.initials: ML
authors.lastname: Leung
authors.affiliation: Department of Genetics, University of Texas MD Anderson Cancer Center, Houston, Texas, USA.
authors.forename: Yong
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Department of Genetics, University of Texas MD Anderson Cancer Center, Houston, Texas, USA.|Graduate Program in Genes and Development, Graduate School of Biomedical Sciences, University of Texas Health Science Center at Houston, Houston, Texas, USA.
authors.forename: Charissa
authors.initials: C
authors.lastname: Kim
authors.affiliation: Department of Genetics, University of Texas MD Anderson Cancer Center, Houston, Texas, USA.
authors.forename: Ruli
authors.initials: R
authors.lastname: Gao
authors.affiliation: Department of Genetics, University of Texas MD Anderson Cancer Center, Houston, Texas, USA.
authors.forename: Jerry
authors.initials: J
authors.lastname: Jiang
authors.affiliation: Department of Genetics, University of Texas MD Anderson Cancer Center, Houston, Texas, USA.
authors.forename: Emi
authors.initials: E
authors.lastname: Sei
authors.affiliation: Department of Genetics, University of Texas MD Anderson Cancer Center, Houston, Texas, USA.|Graduate Program in Genes and Development, Graduate School of Biomedical Sciences, University of Texas Health Science Center at Houston, Houston, Texas, USA.|Department of Bioinformatics and Computational Biology, University of Texas MD Anderson Cancer Center, Houston, Texas, USA.
authors.forename: Nicholas E
authors.initials: NE
authors.lastname: Navin
mesh_terms: D002467:Cell Nucleus; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D017422:Sequence Analysis, DNA / Q000379:methods*; D059010:Single-Cell Analysis / Q000379:methods*; D013997:Time Factors