Identification of polymorphisms by genomic denaturing gradient gel electrophoresis: application to the proximal region of human chromosome 21.
Genomic Denaturing Gradient Gel Electrophoresis (gDGGE) provides an alternative to the standard method of restriction fragment length polymorphism (RFLP) analysis for identifying polymorphic sequence variation in genomic DNA. For gDGGE, genomic DNA is cleaved by restriction enzymes, separated in a polyacrylamide gel containing a gradient of DNA denaturants, and then transferred by electroblotting to nylon membranes. Unlike other applications of DGGE, gDGGE is not limited by the size of the probe and does not require probe sequence information. gDGGE can be used in conjunction with any unique DNA probe. Here we use gDGGE with probes from the proximal region of the long arm of human chromosome 21 to identify polymorphic DNA sequence variation in this segment of the chromosome. Our screening panel consisted of DNA from nine individuals, which was cleaved with five restriction enzymes and submitted to electrophoresis in two denaturing gradient conditions. We detected at least one potential polymorphism for nine of eleven probes that were tested. Two polymorphisms, one at D21S4 and one at D21S90, were characterized in detail. Our study demonstrates that gDGGE is a fast and efficient method for identifying polymorphisms that are useful for genetic linkage analysis.
Department of Physiology, University of California, San Francisco 94143.
M
M
Burmeister
G
G
diSibio
D R
DR
Cox
R M
RM
Myers
D002891:Chromosomes, Human, Pair 21; D015342:DNA Probes; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D009691:Nucleic Acid Denaturation; D009693:Nucleic Acid Hybridization; D012150:Polymorphism, Restriction Fragment Length; D015183:Restriction Mapping
D015342:DNA Probes