Double-stranded DNA analysis by capillary electrophoresis with laser-induced fluorescence using ethidium bromide as an intercalator.
We have developed a rapid and sensitive method to analyze double-stranded DNA by capillary electrophoresis equipped with a laser-induced fluorescence detector. Using ethidium bromide as an intercalator in polyacrylamide gel-filled capillaries along with a green He/Ne laser source (excitation: 543 nm; emission: 600 nm), we found the detection sensitivity for a single DNA-ethidium bromide complex was in the picogram level with a resolution of one base pair separation between two DNA fragments. In the case of the separation of a phi X174 DNA-HaeIII digest, sizes from 72 to 1353 bp were well resolved within 12 min. For the analysis of PCR-amplified DNA, low levels of amplified DNA fragments could be detected that could not be visualized in agarose gel after standard ethidium bromide staining. Considering the high resolving power and sensitivity, we believe that this method can be a useful tool for the analysis of double-stranded DNA especially for PCR-amplified DNA fragments.
Liu
M S
MS
Advanced Technology Center, D-20-A, Beckman Instruments, Fullerton, CA 92634, USA.
Zang
J
J
Evangelista
R A
RA
Rampal
S
S
Chen
F T
FT
D001483:Base Sequence; D002197:Capillary Action; D004247:DNA / Q000235:genetics*; D004586:Electrophoresis / Q000379:methods*; D004996:Ethidium; D005456:Fluorescent Dyes; D007364:Intercalating Agents; D007834:Lasers; D008969:Molecular Sequence Data
D005456:Fluorescent Dyes; D007364:Intercalating Agents; D004247:DNA; D004996:Ethidium