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Characterization of dye‐induced mobility shifts affecting DNA sequencing in poly(ethylene oxide) sieving matrix
Author(s) -
Tan Hongdong,
Yeung Edward S.
Publication year - 1997
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.1150181529
Subject(s) - bodipy , fluorescence , capillary electrophoresis , dna sequencer , rhodamine , dna , ethylene oxide , chemistry , electrophoresis , oxide , rhodamine b , dna sequencing , photochemistry , primer (cosmetics) , analytical chemistry (journal) , materials science , chromatography , polymer , organic chemistry , biochemistry , quantum mechanics , photocatalysis , copolymer , catalysis , physics
The influence of three classes of fluorescence labels including dipyrrometheneboron difluoride (BODIPY), energy transfer (ET) and conventional fluorescein and rhodamine (ABI), on DNA sequencing has been examined with laser‐induced fluorescence detection and poly(ethylene oxide)‐filled capillary electrophoresis. DNA sequencing fragments were generated by dye‐labeled primer cycle‐sequencing reactions in a hot‐air thermal cycler. A parameter, relative‐induced shift, was introduced to quantify the uniformity of electrophoretic mobilities of these fragments. BODIPY was found to have the smallest, but nonzero, effect for dye‐induced nonuniformity. Although ET dyes provided the highest sensitivity due to their unique spectroscopic properties, they were found to lack photostability compared to BODIPY and ABI dyes. Characterization also brings out some important tips for selecting the suitable dye set for the two‐channel ratio‐based DNA base‐calling method.