The basic snRNA-seq method requires 4 main steps: tissue processing, nuclei isolation, cell sorting, and sequencing.
[3] In response, the DroNc-Seq method of massively parallel snRNA-seq with droplet technology was developed by researchers from the Broad Institute of MIT and Harvard.
[3] In this technique, nuclei that have been isolated from their fixed or frozen tissue are encapsulated in droplets with uniquely barcoded beads that are coated with oligonucleotides containing a 30-terminal deoxythymine (dT) stretch.
[4] Sequencing this cDNA produces the transcriptomes of all the single nuclei being looked at and these can be used for many purposes, including identification of unique cell types.
For this reason, compared to scRNA-seq, snRNA-Seq is more appropriate to profile gene expression in cells that are difficult to isolate (e.g. adipocytes, neurons), as well as preserved tissues.
[5] Additionally, the nuclei required for snRNA-seq can be obtained quickly and easily from fresh, lightly fixed, or frozen tissues, whereas isolating single cells for single-cell RNA-seq (scRNA-seq) involves extended incubations and processing.
[8] As snRNA-seq has emerged as an alternative method of assessing a cell's transcriptome through the isolation of single nuclei, it has been possible to conduct single-neuron studies from postmortem human brain tissue.
They found snRNA-seq accomplishes an equivalent gene detection rate to that of scRNA-seq in adult kidney with several significant advantages (including compatibility with frozen samples, reduced dissociation bias and so on ).