Targeting individual cells by barcode in pooled sequence libraries

Nucleic Acids Res. 2019 Jan 10;47(1):e4. doi: 10.1093/nar/gky856.

Abstract

Transcriptional profiling of thousands of single cells in parallel by RNA-seq is now routine. However, due to reliance on pooled library preparation, targeting analysis to particular cells of interest is difficult. Here, we present a multiplexed PCR method for targeted sequencing of select cells from pooled single-cell sequence libraries. We demonstrated this molecular enrichment method on multiple cell types within pooled single-cell RNA-seq libraries produced from primary human blood cells. We show how molecular enrichment can be combined with FACS to efficiently target ultra-rare cell types, such as the recently identified AXL+SIGLEC6+ dendritic cell (AS DC) subset, in order to reduce the required sequencing effort to profile single cells by 100-fold. Our results demonstrate that DNA barcodes identifying cells within pooled sequencing libraries can be used as targets to enrich for specific molecules of interest, for example reads from a set of target cells.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Blood Cells / classification
  • Cell Lineage / genetics
  • DNA / classification
  • DNA / genetics*
  • DNA Barcoding, Taxonomic / methods*
  • High-Throughput Nucleotide Sequencing / methods*
  • Humans
  • Leukocytes, Mononuclear / cytology
  • Multiplex Polymerase Chain Reaction / methods*
  • Single-Cell Analysis / methods

Substances

  • DNA