Table 2.
Determination of Tm values of different neomycin-binding aptamers
| Aptamer | Tm in °Ca 0 µM neomycin | Tm in °Ca 10 µM neomycin | ΔTm | Regulatory Factorb |
|---|---|---|---|---|
| N1-3/0-CUU | 50.5 ± 0.2 | 71.8 ± 0.5 | 21.3 | 3.9 |
| R23 | 57.5 ± 0.5 | 73.1 ± 0.6 | 15.6 | 1.1 |
| N1(R23) | 51.3 ± 0.5 | 71.5 ± 0.4 | 20.2 | 1.6 |
| N1(A) | 54.6 ± 0.5 | 68.3 ± 0.8 | 13.7 | 1.0 |
| N1-3/0-CUC | 50.5 ± 0.2 | 72.5 ± 0.3 | 22.0 | 3.5 |
| N1-2/0-CU | n.d. | 74.0 ± 0.4 | n.d. | 3.7 |
| N1-1/0-C | 61.0 ± 0.5 | 77.1 ± 0.2 | 16.1 | 1.9 |
| N1-1/0-U | 61.6 ± 0.5 | 77.6 ± 0.7 | 16.1 | 1.8 |
| N1-0/0 | 74.2 ± 0.4 | 80.5 ± 0.5 | 6.3 | 1.0 |
| N1-2/1-CU/A | 60.3 ± 0.3 | 71.4 ± 0.5 | 11.1 | 1.1 |
| N1-3/1-CUU/A | 56.0 ± 0.5 | 69.3 ± 0.8 | 13.3 | 1.1 |
n.d.: not determined. The aptamer N1-2/0-CU shows biphasic melting behavior in the absence of neomycin.
aMelting analysis was performed using 1 µM aptamer RNA in 20 mM Na-cacodylate pH 6.8 and 100 mM NaCl. Melting curves were recorded in triplicates. Buffer with or without neomycin was analyzed in parallel as a blank and substracted from all data.
bEfficiency of regulation is given as the ratio of relative fluorescence with and without neomycin.