Quantitation of polymerase chain reaction-amplified DNA fragments by capillary electrophoresis and laser-induced fluorescence detection.
Quantitation of DNA fragments amplified by polymerase chain reaction (PCR) is needed for the determination of target DNA in molecular biology. Capillary electrophoresis in entangled polymer solution coupled to laser-induced fluorescence detection was assessed as an alternative technique to conventional slab gel methods to monitor competitive PCR, which consists of amplifying an internal standard fragment under the same conditions as the target fragment. The fluorescence signal was generated either through end-labeling of the fragments using 5'-fluorescein-labeled primers or through intercalation of ethidium bromide along the double strand. It is shown that the more accurate and reliable results were obtained using this latter pathway.
Vincent
U
U
Service des Biotechnologies, CEB, Vert-le-Petit, France.
Patra
G
G
Therasse
J
J
Gareil
P
P
D001408:Bacillus anthracis / Q000235:genetics* / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D004269:DNA, Bacterial / Q000032:analysis*; D019075:Electrophoresis, Capillary / Q000379:methods*; D005452:Fluoresceins; D005456:Fluorescent Dyes; D007834:Lasers; D008562:Membrane Glycoproteins; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D005452:Fluoresceins; D005456:Fluorescent Dyes; D008562:Membrane Glycoproteins; C055764:S-layer proteins