Keith C. Backman

Ligase Chain Reaction

Ligase chain reaction (LCR), employing just oligonucleotide probes and Principle and DNA ligase, is capable of detecting approximately <_ 1000 copies of a specific applications target DNA sequence in the presence of a vast excess of other DNA sequence information. Since the first description in 1989 (Backman and Wang, 1989; Royer et al., 1989; Wallace, 1989; Wu and Wallace, 1989; Orgel, 1989; Richards and Jones, 1989) LCR has been improved by the employment of a thermostable DNA ligase in conjunction with nonradioactive detection (Bond et al., 1990). The ability of LCR to distinguish between normal βAand sickel βS-globin genotypes from 10 µl of whole blood using probes having a mismatch or full complementarity at the point of ligation has recently been reported (Barany, 1991), demonstrating the potential of LCR in allele-specific detection.

Enhanced Signal Generation by Target Amplification

The polymerase chain reaction (PCR) represents the most common and widespread method for the direct amplification of specific sequences of nucleic acid target molecules. The basic reaction is comprised of three steps: 1. Denaturation of the target DNA 2. Annealing of sequence specific primers 3. Template-specific elongation of these primers with a DNA polymerase and desoxynucleotides