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Separation of long DNA fragments by capillary gel electrophoresis with laser‐induced fluorescence detection
Author(s) -
Nishikawa Tetsuo,
Kambara Hideki
Publication year - 1994
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.1150150136
Subject(s) - capillary electrophoresis , detection limit , resolution (logic) , laser induced fluorescence , analytical chemistry (journal) , electrophoresis , chemistry , chromatography , fluorescence , capillary action , electric field , laser , gel electrophoresis , materials science , optics , physics , quantum mechanics , artificial intelligence , computer science , composite material , biochemistry
Long DNA fragments were separated by capillary gel electrophoresis with laser‐induced fluorescence detection, and two approaches to obtaining high‐resolution separation of DNA fragments were examined. One method achieved high resolution at the expense of detection time, while the other achieved both high resolution and fast detection. The former approach used a 300 cm long capillary with a gel concentration of 4% T and an electric field strength of 70 V/cm. The resolution limit in this electrophoresis was 800 bases with a resolution value of 0.5 for adjacent peaks. Under these conditions, the measurement time was 44 h. The latter approach used a 200 cm long capillary with a gel concentration of 3% T and an electric field strength of 170 V/cm. The resolution limit in this electrophoresis was 680 bases with a resolution value of 0.5 for adjacent peaks, and the measurement time was only 10 h. With both approaches, fragments with less than 600 bases were efficiently separated. The resolution values for adjacent peaks with less than 500 bases are greater than 1.0, and those for peaks with less than 100 bases are greater than 2.7. These approaches thus improve the accuracy of the base sequence determination.