ELECTROPHORETIC SEPARATIONS OF POLYMERASE CHAIN REACTION-AMPLIFIED DNA FRAGMENTS IN DNA TYPING USING A CAPILLARY ELECTROPHORESIS LASER-INDUCED FLUORESCENCE SYSTEM

Analysis of polymerase chain reaction (PCR)-amplified DNA fragments fo r human identification requires high-resolution separation and efficie nt detection of amplified alleles. Capillary electrophoresis (CE) with its speed, automation, high resolution and efficiency shows promise f or analyzing the amplified DNA fragments. CE with UV detection, howeve r, suffers from lack of detector sensitivity owing to the limited dete ction path length of the capillary. By the use of intercalating dyes ( TOTO and YOYO) a laser-induced fluorescence (LIF) detection system can provide much greater sensitivity for detecting DNA fragments. Femtogr am quantities of dsDNA (Phi X174 HaeIII restriction digest mixture) pe r nanoliter of injected volume have been detected. Application of CE-L IF to analysis of PCR-amplified DNA fragments from three different gen etic loci (apolipoprotein B, VNTR locus D1S80, mitochondrial DNA) is s hown here. Further, the resolving power of a polymer-network capillary separation system is compared to that of a capillary-gel separation s ystem.