| Abstract
| - A continuous segmented-flow method for sequential DNAamplification is described in order to provide a basis forhigh-throughput genetic analysis. The approach allows animmediate distinction between amplified and nonamplified products. A mixture of sample and reagents areloaded in the form of small segments one after another ina 15-m-long narrow-bore Teflon tube, coiled such as tobe repeatedly exposed to three different temperaturezones. After having passed the heated zones, the samplesare mixed with an intercalating dye by flow injection andsequentially detected on-line by laser-induced fluorescence. The aqueous samples travel as separate segmentsin a continuous flow of an immiscible, organic liquid.Perfluorodecalin was shown to be particularly suitabledue to its hydrophobicity and inert properties. To reducecarryover between samples, an intermediate water plugbetween two consecutive samples was required. Selectedregions from human genomic DNA were successfullyamplified in 300-nL volumes after 30 passes through theheated zones. The total reaction time was ∼45 min, andthe detection interval between individual samples was 1min. Automation and the possibility to further reducesample volumes, as well as to employ many reactioncolumns simultaneously, should provide a platform foran extremely high throughput.
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