High resolution mapping using fluorescence in situ hybridization to extended DNA fibers prepared from agarose-embedded cells.
Fluorescence in situ hybridization (FISH) and pulse field gel electrophoresis (PFGE) are essential techniques in physical mapping and in positional cloning. We present a technique that utilizes agarose-embedded high molecular weight DNA prepared for PFGE as a target for FISH. The agarose blocks are melted, and the DNA is extended on a poly-L-lysine-coated microscope slide. The resulting DNA fibers appear on the slide as long straight strands and are a suitable target for high resolution FISH mapping as demonstrated here with cosmid and plasmid hybridizations.
University of Helsinki, Finland.
M
M
Heiskanen
R
R
Karhu
E
E
Hellsten
L
L
Peltonen
O P
OP
Kallioniemi
A
A
Palotie
D002874:Chromosome Mapping / Q000379:methods*; D003360:Cosmids; D004247:DNA / Q000378:metabolism*; D016521:Electrophoresis, Gel, Pulsed-Field; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D008214:Lymphocytes / Q000737:chemistry; D008856:Microscopy, Fluorescence; D010957:Plasmids; D011107:Polylysine; D015183:Restriction Mapping; D012685:Sepharose
D011107:Polylysine; D004247:DNA; D012685:Sepharose