Multiplexed optical barcoding and sequencing for spatial omics.

Spatial omics has brought a fundamental change in the way that we study cell and tissue biology in health and disease. Among various spatial omics methods, genome-scale imaging allows transcriptomic, 3D-genomic, and epigenomic profiling of individual cells with high spatial (subcellular) resolution but typically requires a preselection of targeted genes or genomic loci. On the other hand, spatially dependent barcoding of molecules followed by sequencing provides untargeted, genome-wide profiling but has a lower spatial resolution than imaging-based methods. Here, we report a spatial omics method that could potentially combine the power of the two approaches using optically controlled spatial barcoding followed by sequencing. Specifically, we utilize patterned light to encode the locations of molecules in tissues using oligonucleotide-based barcodes and then identify the barcoded molecular content, such as mRNAs, by sequencing. This optical barcoding method is designed with multiplexing and error-correction capability and achieved by a light-directed ligation chemistry that attaches distinct nucleic-acid sequences to the reverse transcribed cDNA products at different locations. As a proof of principle for this method, we demonstrated high-efficiency in situ light-directed ligation, spatially dependent barcoding with multiplexed light-controlled ligations at the single-cell level, and high-accuracy detection of spatially barcoded mRNAs in cells.