Xdrop: Targeted sequencing of long DNA molecules from low input samples using droplet sorting

Area of Science:

• Genomics
• Molecular Biology
• Biotechnology

Background:

• Short-read sequencing technologies have limitations in resolving complex genomic regions.
• Long-read sequencing offers advantages for genome gap closure, structural variation analysis, and repetitive element sequencing.
• Targeted enrichment of long DNA molecules is crucial for studying specific genomic loci.

Purpose of the Study:

• To introduce and evaluate Xdrop, a novel microfluidic-based system for targeted enrichment of long DNA molecules.
• To demonstrate the utility of Xdrop for analyzing complex genomic structures and mutations.
• To showcase Xdrop's capability in enriching specific genomic regions for downstream sequencing.

Main Methods:

• Xdrop technology utilizes microfluidics to isolate long DNA fragments within millions of droplets.
• Targeted DNA sequences are fluorescently labeled within droplets and sorted using flow cytometry.
• Enriched long DNA molecules are then analyzed using sequencing technologies, such as PacBio.

Main Results:

• Xdrop successfully enriched human papillomavirus 18 integrated into HeLa cell genomes, resolving integrations at base-pair resolution.
• Captured fragments extended up to 30 kb into the human genome at integration sites.
• The TP53 locus was enriched in a leukemia cell line, enabling successful phasing of coexisting mutations using PacBio sequencing.

Conclusions:

• Xdrop is an efficient and effective technology for the targeted enrichment of long DNA molecules.
• The system facilitates the study of complex genomic regions, including viral integrations and phased mutations.
• Xdrop provides a valuable tool for advancing genomic research and diagnostics.