Abstract
Molecular single cell analyses provide insights into physiological and pathological processes. Here, in a stepwise approach, we first evaluate 19 protocols for single cell small RNA sequencing on MCF7 cells spiked with 1pg of 1,006 miRNAs. Second, we analyze MCF7 single cell equivalents of the eight best protocols. Third, we sequence single cells from eight different cell lines and 67 circulating ...
Abstract
Molecular single cell analyses provide insights into physiological and pathological processes. Here, in a stepwise approach, we first evaluate 19 protocols for single cell small RNA sequencing on MCF7 cells spiked with 1pg of 1,006 miRNAs. Second, we analyze MCF7 single cell equivalents of the eight best protocols. Third, we sequence single cells from eight different cell lines and 67 circulating tumor cells (CTCs) from seven SCLC patients. Altogether, we analyze 244 different samples. We observe high reproducibility within protocols and reads covered a broad spectrum of RNAs. For the 67 CTCs, we detect a median of 68 miRNAs, with 10 miRNAs being expressed in 90% of tested cells. Enrichment analysis suggested the lung as the most likely organ of origin and enrichment of cancer-related categories. Even the identification of non-annotated candidate miRNAs was feasible, underlining the potential of single cell small RNA sequencing. Technologies for small non-coding RNA sequencing at the single-cell level are less mature than for sequencing mRNAs. Here the authors evaluate available protocols for analysis of circulating lung cancer tumour cells.
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