Table 2.
Comparison of T m values for replication relevant DNA architectures containing individual 3′ and 5′ overhangs, a terminal phosphate, and presence of both an overhang and phosphate. The presence of a 3′ or 5′ phosphate (PO4), hydroxyl (OH) and a 6 mer overhang is indicated as an “X” or “Y” at the end of the core dsDNA sequences. ΔT m values were obtained by subtracting the T m of each construct from the T m value of the core duplex.
| ID | Base Sequence | T m (°C) | ΔT m (°C) | |
|---|---|---|---|---|
| 1 | 5′-CGCAACCTGAAAX | |||
| 3′-GCGTTGGACTTTY | ||||
| 1G | X = OH | Y = OH | 51.7 ± 0.2 | — |
| 1H | X = OH | Y = PO4 | 51.3 ± 0.0 | −0.4a |
| 1I | X = AAAAAA | Y = OH | 48.9 ± 0.3 | −2.8b |
| 1J | X = AAAAAA | Y = PO4 | 47.9 ± 0.1 | −3.4b |
| −1.0a | ||||
| 1K | X = GACTGT | Y = OH | 49.8 ± 0.3 | −1.9b |
| 1L | X = GACTGT | Y = PO4 | 48.7 ± 0.2 | −2.6b |
| −1.1a | ||||
|
| ||||
| 2 | 5′-GCGTTGGACTTTX | |||
| 3′-CGCAACCTGAAAY | ||||
| 2G | X = OH | Y = OH | 52.9 ± 0.2 | — |
| 2H | X = PO4 | Y = OH | 52.0 ± 0.2 | −0.9a |
| 2I | X = OH | Y = AAAAAA | 51.8 ± 0.1 | −1.1b |
| 2J | X = PO4 | Y = AAAAAA | 50.5 ± 0.3 | −1.5b |
| −1.3a | ||||
| 2K | X = OH | Y = GACTGT | 51.6 ± 0.2 | −1.3b |
| 2L | X = PO4 | Y = GACTGT | 50.4 ± 0.2 | −1.6b |
| −1.2a | ||||
aIndicates the ΔT m introduced by an individual phosphate. ΔT m was determined by subtracting substrate G from H, I from J and K from L. bIndicates the ΔT m introduced by an individual overhang. ΔT m was determined by subtracting substrate G from I, G from K, H from J, and H from L.