Table 1. Homozygous and heterozygous templates and their photocleavage.
| SNP alleles present* | Mismatches generated† | Rhphzi photocleavage‡ | Rhchrysi photocleavage‡ |
|---|---|---|---|
| AT | None | ||
| TA | None | ||
| GC | None | ||
| CG | None | ||
| AT + TA | AA + TT | X | |
| AT + GC | AC + GT | X | X |
| AT + CG | AG + CT | ||
| TA + GC | TC + GA | X | X |
| TA + CG | TG + CA | X | X |
| GC + CG | GG + CC | X | X |
*
The base pair(s) at the polymorphic site present in the template plasmid. The first base indicates the template plasmid used.
†
Upon denaturing and annealing of the PCR-amplified template, mismatches can be generated as indicated if heterozygousity exists. DNA duplexes are denatured in the presence of 10 mM Tris (pH 8.5) and annealed with the addition of buffer to make a final concentration of 60 mM Tris·HCl (pH 7.5)/10 mM MgCl2/100 mM NaCl/1 mM dithioerythritol. This buffer is diluted to half with the addition of metal complex to 500 nM Rhchrysi or 200 nM Rhphzi.
‡
Both Rhphzi and Rhchrysi generate site-specific cleavage products, upon irradiation at 340 nm for 1 h or 442 nm for 1 h, respectively, as denoted by X.