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TARGET-Seq: A Protocol for High-Sensitivity Single-Cell Mutational Analysis and Parallel RNA Sequencing.

ABSTRACT: Single-cell RNA-sequencing technologies are ideally placed to resolve intratumoral heterogeneity. However, the lack of coverage across key mutation hotspots has precluded the correlation of genetic and transcriptional readouts from the same single cell. To overcome this, we developed TARGET-seq, a protocol for TARGETed high-sensitivity single-cell mutational analysis with extremely low allelic dropout rates, parallel RNA SEQuencing, and cell-surface proteomics. Here, we present a detailed step-by-step protocol for TARGET-seq, including troubleshooting tips, approaches for automation, and methods for high-throughput multiplexing of libraries. For complete details on the use and execution of this protocol, please refer to Rodriguez-Meira et al. (2019).

SUBMITTER: Rodriguez-Meira A

PROVIDER: S-EPMC7757016 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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TARGET-Seq: A Protocol for High-Sensitivity Single-Cell Mutational Analysis and Parallel RNA Sequencing.

Rodriguez-Meira Alba A O'Sullivan Jennifer J Rahman Haseeb H Mead Adam J AJ

STAR protocols 20201021 3

Single-cell RNA-sequencing technologies are ideally placed to resolve intratumoral heterogeneity. However, the lack of coverage across key mutation hotspots has precluded the correlation of genetic and transcriptional readouts from the same single cell. To overcome this, we developed TARGET-seq, a protocol for TARGETed high-sensitivity single-cell mutational analysis with extremely low allelic dropout rates, parallel RNA SEQuencing, and cell-surface proteomics. Here, we present a detailed step-b ...[more]

PMID: 33377019

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