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Assays & screens

In situ RNA sequencing in fixed cells and tissues

In situ RNA sequencing could help reveal genetic heterogeneity in complex samples for clinical diagnostics. In fresh-frozen cells or tissues, single-cell RNA sequence information was derived from sequencing by ligation after padlock probing and rolling-circle amplification. Control in situ experiments on the human b-actin transcript showed an average accuracy of 98.6%. In sections of frozen tissue from breast cancer biopsies, 39 probes, including 21 from a prognostic gene panel, were combined in a single multiplex reaction, and the results were validated by comparison to published RNA-sequencing data.
Next steps could include optimization of the workflow for handling large numbers of samples and comparison of this method directly with bulk expression profiling for prognostic evaluation.

SciBX 6(31); doi:10.1038/scibx.2013.834
Published online Aug. 15, 2013

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Ke, R. et al. Nat. Methods; published online July 14, 2013;
Contact: Mats Nilsson, Stockholm University, Stockholm, Sweden
Contact: Carolina Wählby, Broad Institute of MIT and Harvard, Cambridge, Mass.