SPRITE

SPRITE maps genome-wide higher-order three-dimensional interactions among DNA, RNA, and proteins in the nucleus using iterative split-and-pool barcoding to enable reconstruction of spatial contacts.

Key Features:

• Cross-linking: Covalent cross-linking of interacting DNA, RNA, and protein molecules preserves native nuclear complexes for downstream analysis.
• Split-and-pool barcoding: Iterative split-and-pool barcoding across 96-well plates with ligation of unique tag sequences ensures components of an original complex acquire identical barcode sequences.
• Sequencing of barcoded fragments: High-throughput sequencing of DNA fragments with associated barcodes produces reads that encode complex membership.
• Barcode-based interaction reconstruction: Reads sharing identical barcodes are matched to reconstruct pairwise and higher-order contacts, including contacts involving thousands of molecules.
• Automated computational pipeline: An automated pipeline converts raw fastq files into SPRITE interaction matrices for downstream analysis.

Scientific Applications:

• Genome architecture mapping: Genome-wide identification of pairwise and higher-order chromatin contacts to define nuclear spatial organization.
• Chromatin–gene regulation studies: Investigation of how DNA packaging and higher-order interactions influence gene expression across cell types.
• RNA and protein association analysis: Characterization of RNA and protein molecules associated with DNA within nuclear complexes.
• Disease and functional genomics: Examination of chromatin architecture alterations relevant to cellular function and disease mechanisms.

Methodology:

High-throughput sequencing of barcoded DNA fragments followed by matching reads with identical barcodes to reconstruct spatial interactions and automated generation of SPRITE interaction matrices from raw fastq files.