SenNet URMC 10X Genomics Single-Nucleus RNA-Sequencing for Transcriptomic Profiling v1

10X Genomics Single Cell 3' (v3.1) RNA sequencing is a microdroplet-based method that permits the effective capture and sequencing of the mRNA and pre-mRNA molecules from single nuclei [1]. RNA molecules are transcribed and processed within the nucleus before exporting to ER for translation into proteins. As such, nuclear RNA is a mixture of nascent transcripts, partially or fully processed mRNA, and various non-coding RNA molecules. The total RNA content within the nucleus is roughly 10% of the RNA content in a whole cell, but has been found to accurately represent whole cell expression values in adult human tissues [2,3] including the kidney [4]. Nuclei can be readily isolated from frozen tissues with a combination of chemical and physical treatments that can effectively circumvent the non-uniform or incomplete dissociation of solid tissues into single cells, as well as RNA degradation or artefacts (such as stress response) during dissociation. Here we present a modified version of the published 10X protocol [1] that we have adapted for the processing of adult human kidney nuclei. References 1. Chromium Next GEM Single Cell 3’ Reagent Kits v3.1(Dual Index) User Guide. Document Number: CG000315. October 2022. 2. Lake et al. (2016). Science, doi:10.1126/science.aaf1204. 3. Lake et al. (2018). Nature Biotechnology, doi:10.1038/nbt.4038. 4. Lake et al. (2019). Nature Communications, doi:10.1038/s41467-019-10861-2.