Polymerase chain reaction heteroduplex polymorphism analysis by entangled solution capillary electrophoresis.
Heteroduplex DNA polymorphism analysis (HPA) makes use of conformational polymorphisms to alter electrophoretic mobility of fragments and can be used to detect non-restrictable loci. We have developed a novel application of entangled solution capillary electrophoresis (ESCE) to separate heteroduplex and homoduplex DNA molecules. The addition of ethidium bromide and glycerol to the free solution sieving buffer resulted in the improved peak resolution and good reproducibility. Reannealed polymerase chain reaction products could be used directly for mutation screening and with fully automated ESCE the entire HPA may be completed in less than 30 min including sample handling. This technology could provide a rapid and highly efficient way for screening rare mutations among large numbers of individuals.
Department of Molecular and Cell Biology, University of Aberdeen, UK.
J
J
Cheng
T
T
Kasuga
K R
KR
Mitchelson
E R
ER
Lightly
N D
ND
Watson
W J
WJ
Martin
D
D
Atkinson
D001483:Base Sequence; D002021:Buffers; D004271:DNA, Fungal / Q000032:analysis*; D004586:Electrophoresis; D004996:Ethidium; D008969:Molecular Sequence Data; D009692:Nucleic Acid Heteroduplexes / Q000032:analysis*; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic
D002021:Buffers; D004271:DNA, Fungal; D009692:Nucleic Acid Heteroduplexes; D004996:Ethidium