High-throughput single nucleus total RNA sequencing of formalin-fixed paraffin-embedded (FFPE) tissues by snRandom-seq. Mus musculus

Formalin-fixed paraffin-embedded (FFPE) tissues constitute a vast and valuable patient material bank. It is still challenging to achieve single cell/nucleus RNA (sc/snRNA) profile in FFPE tissues. Here, we developed a droplet-based snRNA sequencing technology (snRandom-seq) for FFPE tissues by capturing full-length total RNAs with random primers. snRandom-seq showed a minor doublet rate (0.3%), a much higher RNA coverage, and detected more non-coding RNAs and nascent RNAs, compared with state-of-art high-throughput scRNA-seq technologies. We used snRandom-seq to profile ~20,000 single nuclei from multiple FFPE mouse tissues. A median of >3,000 genes per nucleus was detected and 25 typical cell types were identified. Moreover, we applied snRandom-seq on a clinical FFPE human liver cancer specimen and revealed an interesting subpopulation of nuclei with high proliferative activity. Our method provides a powerful snRNA-seq platform for clinical FFPE specimens and promises enormous applications in biomedical research.