Single nuclei RNA sequencing (snRNA-seq) using Evercode combinatorial barcoding is a powerful tool to capture whole transcriptomes of up to 1 million nuclei from up to 96 samples to identify both known and novel gene expression profiles. The power of snRNA-seq is the ability to detect thousands of differential genes to capture cellular heterogeneity which is important for very complex tissues such as the mammalian brain where there are a myriad of cell types within brain structures, many defined by their specific cell-cell interactions and location.
Since traditional snRNA-seq sacrifices spatial information due to the dissociation of samples into single nuclei suspensions we sought to complement Evercode snRNA-seq with spatial RNAscope RNA in situ hybridization (RNA-ISH). RNA-ISH provides detailed spatial localization of a set of user defined genes at single cell resolution. We generated a set of RNAscope probes to both canonical genes from literature and ones found by snRNA-seq differential gene expression (DGE) to validate, determine colocalization, and visualize spatial distribution of these targets. We observed a remarkable concordance of gene expression between snRNA-seq and RNA-ISH, exhibiting both cell specificity and brain region specificity, highlighting the power of these complementary techniques to gain a holistic understanding of biological complexity
