Automated DNA detection with fluorescence-based technologies

Abstract

Abstract The use of fluorescence-based technologies in DNA detection systems has truly come of age. Automated DNA sequencing instruments, which were once relegated to core sequencing facilities at a few large institutions, are now found at thousands of laboratories throughout the world. The capabilities of these instruments have been expanded to encompass new technical develop ments such as microsatellites (1,2), random amplified polymorphic DNA (RAPD) markers (3,4), variable number tandem repeats (VNTRs; 5) and single-strand conformational polymorphism (SSCP; 6). The amenability of this technology to automation has made it especially useful in large-scale sequencing and genotyping projects; however, high sample throughput can be accomplished without expensive robotic systems. DNA storage, polymerase chain reaction (PCR) amplification, DNA sequence reactions, and sample loading can all be accomplished in a format compatible with a standard 96-well plate. This allows for the use of multi-channel pipettes to facilitate sample processing and multi-channel syringes for sample loading. This chapter will assess current applications of fluorescence-based DNA detection systems with emphasis on the two most widely used applications, DNA sequence analysis and microsatellite genotyping.