A 3-WAVELENGTH LABELING APPROACH FOR DNA-SEQUENCING USING ENERGY-TRANSFER PRIMERS AND CAPILLARY-ELECTROPHORESIS

Capillary electrophoresis DNA sequencing has been accomplished by usin g four different energy transfer primers and three fluorescence detect ion channels. Methods have also been developed to deconvolve the three -color data into the four base concentrations. The nonnegative least s quares and model selection method resulted in the best accuracy. The t hree-color data were compared to sequencing data obtained using four d etection channels and four energy transfer primers. The average accura cy rates obtained over three 500 base M13mp18 runs using three-color c oding were 96% including 18 uncallable compressions and 99.6% if these compressions are excluded. The average accuracy rate obtained using f our-color coding was 96.3% including 18 uncallable compressions and 99 .9% if these compressions are excluded. Although it is unlikely that t hree-color schemes will replace four-color sequencing, these methods h ave exposed basic concepts that will be useful in the development of h igher-order multiplex coding methods for DNA analysis.