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Updated: Jun 13, 2025
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An Ultrahigh-throughput Microfluidic Platform for Single-cell Genome Sequencing
Published on: May 23, 2018
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Protocol for high-quality RNA sequencing, cell surface protein analysis, and genotyping in single cells using
N Asger Jakobsen1, Sven Turkalj1, Paresh Vyas2
1MRC Molecular Haematology Unit, MRC Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK; Oxford Centre for Haematology, Oxford University Hospitals NHS Foundation Trust, Oxford, UK.
STAR Protocols
|May 23, 2025
Summary
TARGET-seq+ enhances single-cell analysis for cancer research by combining RNA sequencing, protein expression, and genotyping. This optimized method improves sensitivity for studying somatic mutations in pre-malignant and cancerous tissues.
Area of Science:
- Genomics
- Molecular Biology
- Cancer Research
Background:
- Studying somatic mutations in pre-malignant and cancerous tissues is difficult due to noisy single-cell transcriptome data and challenges in identifying cell clonal identity.
- Existing methods lack the sensitivity and multi-omic capabilities required for comprehensive analysis.
Purpose of the Study:
- To optimize and validate TARGET-seq+ for enhanced single-cell analysis.
- To combine RNA sequencing (RNA-seq), cell surface protein expression, and genotyping in single cells with improved sensitivity.
- To provide a detailed protocol and analysis guidance for researchers.
Main Methods:
- Optimization of the TARGET-seq protocol to TARGET-seq+.
- Integration of single-cell RNA sequencing (scRNA-seq), cell surface protein analysis, and genotyping.
- Development of protocols for cell isolation, library preparation, and sequencing.
- Guidance on data analysis, including transcriptome pre-processing and data integration.
Main Results:
- TARGET-seq+ demonstrates improved sensitivity for multi-omic single-cell analysis.
- The protocol enables simultaneous assessment of transcriptome, proteome, and genotype at the single-cell level.
- Successful application in studying somatic mutations in pre-malignant and cancerous tissues.
Conclusions:
- TARGET-seq+ is a powerful, optimized tool for high-sensitivity, multi-omic single-cell analysis.
- This method overcomes previous limitations in studying cellular heterogeneity and somatic mutations.
- The provided protocol facilitates the application of TARGET-seq+ in cancer research and beyond.
