Generation of Paint Probes from Flow-Sorted and Microdissected Chromosomes
FISH with whole chromosome or region-specific painting probes made from either flow-sorted or microdissected chromosomes has revolutionized cytogenetics. Generation of paints from flow-sorted chromosomes relies on the use of an expensive and sophisticated fluorescence-activated cell sorter and suspensions of freshly prepared chromosomes. Preparation of paints from microdissected materials requires an inverted microscope with appropriate micromanipulators and metaphase chromosome spreads on coverslips. Painting probes made from flow-sorted chromosomes generally have better chromosomal coverage and can be used in a wide range of applications between distantly related species, while region-specific probes from microdissection enable higher-resolution analyses restricted to comparative painting between closely related species. Here we provide detailed protocols on generation probes from both flow-sorted and microdissected chromosomes.
KeywordsProbe for FISH Chromosome painting Flow sorting Microdissection Human chromosome-specific paints Animal chromosome-specific paints
While writing this protocol, VT was supported by Budget Projects 0310-2014-0003, 0310-2014-0008, 0310-2014-0009, and DFG. FY, BLN, and NPC are supported by the Wellcome Trust (Grant number WT098051).