MATRIX-ASSISTED LASER DESORPTION IONIZATION FOR SEQUENCING SINGLE-STRANDED AND DOUBLE-STRANDED DNA/

The DNA sequence of a single-stranded and double-stranded template was determined. The templates were sequenced using the chain termination method and cycle sequencing method and detected by matrix-assisted las er desorption/ionization time-of-flight mass spectrometry. The sequenc ing products were analyzed successfully without the laborious and expe nsive methods for removal of the template. Direct sequencing of the do uble-stranded template was achieved with minimal post-reaction purific ations, which could be extremely important for mutation analysis and c linical diagnosis. A systematic study of the mechanisms and kinetics o f sequencing reactions was also performed. The details of this analysi s and directions for future improvements of the quality of sequencing are presented. (C) 1997 by John Wiley & Sons, Ltd.