Table 1. Comparison of the properties of the PCR amplifications with various D primers containing or not a 3′-LNA.
| Regular primers | 3′-LNA primers | Mismatch | |||||
|---|---|---|---|---|---|---|---|
| ΔCtMIS−MAT | E | Bgd (%) | ΔCtMIS−MAT | E | Bgd (%) | ||
| U2− Unmet | 7.2 | 0.79 | 1.5 | 12 | 0.78 | 0.10 | A:C |
| U3− Unmet | 8.8 | 0.75 | 0.7 | 13.3 | 0.76 | 0.05 | A:C |
| L2+ Unmet | 6.8 | 0.72 | 2.5 | 14.4 | 0.66 | 0.07 | A:C |
| L2+ Met | N/A | N/A | N/A | 11.7 | 0.78 | 0.12 | G:T |
| L3− Unmet | 4.5 | 0.76 | 7.9 | 14 | 0.72 | 0.05 | T:G |
| L3− Met (56°C) | 2.6 | 0.71 | 24.8 | 9.5 | 0.79 | 0.40 | C:A |
| L3− Met (58°C) | 4.3 | 0.65 | 11.6 | 10.3 | 0.76 | 0.30 | C:A |
The ΔCtMIS−MAT were determined for each D primer sets with identical amounts of MIS and MAT templates, the efficiency (E) of PCR amplification was determined using standard curves of MIS and MAT templates as described in Figure 2 and the Bgd value amplification with MIS template was calculated using Equation 1. The nature of the mismatch involving the 3′-most base of the D primer and the MIS template is indicated (primer:template). A primer annealing temperature of 56°C was used for all primer sets except for the L3− Met primer set that was analyzed at two temperatures as indicated.