Light-Seq

Light-Seq enables multiplexed spatial indexing of intact biological samples by light-directed DNA barcoding for ex situ full-transcriptome sequencing while preserving tissue spatial context.

Key Features:

• Light-Directed DNA Barcoding: Employs light-directed photocrosslinking of DNA barcodes onto complementary DNAs within intact fixed tissue samples to spatially tag transcripts.
• Spatially Indexed Sequencing Libraries: Uses a one-step DNA stitching reaction to produce pooled sequencing libraries that retain spatial indexes.
• Full-Transcriptome Sequencing of Fixed Samples: Performs ex situ full-transcriptome sequencing on barcoded material from fixed cells and tissues without cellular dissociation.
• Preservation of Sample Integrity: Avoids cellular dissociation, preserving tissue architecture and allowing subsequent analyses after sequencing.
• Integration with Imaging and Staining: Combines in situ imaging and protein staining with next-generation sequencing to correlate transcriptomic data with cellular morphology and protein expression.

Scientific Applications:

• Spatially Resolved Tissue Transcriptomics: Enables spatially resolved analysis of complex tissues, demonstrated by identification of thousands of differentially enriched transcripts across three cellular layers in mouse retinal sections.
• Rare Cell-Type Biomarker Discovery: Facilitates discovery of biomarkers for rare neuronal subtypes, exemplified by identification of markers for dopaminergic amacrine cells from as few as four to eight individual cells per section.

Methodology:

Light-directed photocrosslinking of DNA barcodes onto complementary DNAs; a one-step DNA stitching reaction to stitch barcoded DNAs into pooled spatially indexed libraries; and ex situ sequencing of those libraries.