Targeting individual cells by barcode in pooled sequence libraries.

Area of Science:

• Single-cell genomics
• Molecular biology
• Immunology

Background:

• Single-cell RNA sequencing (scRNA-seq) enables transcriptional profiling of numerous cells simultaneously.
• Analyzing specific cell populations within pooled scRNA-seq libraries is challenging due to library preparation methods.

Purpose of the Study:

• To develop a method for targeted sequencing of selected cells from pooled scRNA-seq libraries.
• To demonstrate the utility of this method for enriching ultra-rare cell types.

Main Methods:

• A multiplexed PCR-based molecular enrichment technique was developed.
• This method targets DNA barcodes within pooled single-cell libraries.
• Enrichment was combined with Fluorescence-Activated Cell Sorting (FACS) for rare cell targeting.

Main Results:

• The method successfully enriched specific cell types from pooled human blood cell scRNA-seq libraries.
• Combining molecular enrichment with FACS enabled efficient targeting of ultra-rare AXL+SIGLEC6+ dendritic cells (AS DCs).
• Sequencing effort for profiling single cells was reduced by 100-fold for rare cell types.

Conclusions:

• DNA barcodes in pooled libraries can serve as targets for molecular enrichment.
• This approach significantly enhances the efficiency of single-cell analysis, particularly for rare populations.
• The method offers a powerful tool for deep profiling of specific cell subsets in complex biological samples.