FiberFISH

FiberFISH, short for DNA fiber FISH, is a cytogenetic technique that maps physical order and spacing of DNA segments along a single stretched DNA molecule by applying fluorescently labeled DNA probes to elongated DNA fibers immobilized on slides. It enables high-resolution mapping of genomic structure beyond conventional metaphase FISH.

In practice, DNA is extracted under gentle conditions to preserve long strands, stretched on silanized glass slides by molecular combing or flow-stretching, fixed, and denatured. Probes corresponding to target loci are hybridized, and bound probes are visualized by fluorescence microscopy. The color pattern along a fiber reveals the linear order and spacing between targets, enabling estimation of distances in kilobases to megabases.

Applications include characterization of structural variation such as duplications, deletions, inversions, and translocations, organization of gene clusters, and validation of assemblies or clone-based maps (BAC/fosmid), particularly in complex or repetitive regions. It is often used in conjunction with other methods like conventional FISH, comparative genomic hybridization, or sequencing to resolve rearrangements.

Advantages and limitations: high resolution on single molecules; can reveal long-range organization; but it is labor-intensive, requires high-quality DNA, relatively low throughput, and interpretation can be challenging in highly repetitive regions; precision depends on fiber extension uniformity and probe spacing.