On-channel base stacking in microchip capillary gel electrophoresis for high-sensitivity DNA fragment analysis.
We evaluated a novel strategy for high-sensitivity DNA fragment analysis in a conventional glass double-T microfluidic chip. The microchip allows for a DNA on-channel concentration based on base stacking (BS) with a microchip capillary gel electrophoretic (MCGE) separation step in a poly(vinylpyrrolidone) (PVP) sieving matrix. Depending if low conductivity caused a neutralization reaction between the hydroxide ions and the run buffer component Tris+, the stacking of DNA fragments were processed in the microchip. Compared to a conventional MCGE separation with a normal electrokinetic injection, the peak heights of 50-2650-base pair (bp) DNA fragments on the MCGE-BS separation were increased 3.9-8.0-fold. When we applied the MCGE-BS method to the analysis of a clinical sample of bovine theileria after PCR reaction, the peak height intensity of the amplified 816-bp DNA fragment from the 18S rRNA of T. buffeli was enhanced 7.0-fold compared to that of the normal injection method.
Kim
Dae-Kwang
DK
Department of Chemistry, Chonbuk National University, Jeonju 561-756, South Korea.
Kang
Seong Ho
SH
D004247:DNA / Q000032:analysis* / Q000737:chemistry; D046209:Electrophoresis, Microchip; D012680:Sensitivity and Specificity
D004247:DNA