rNTP
Dissolve each NTP (ribonucleoside triphosphates) in H2O at an approximate concentration of 100 mM. Use 0.05 M Tris base (or 0.1 N NaOH as appropriate) and a micropipette to adjust the pH of each of the solutions to 7.0 (use pH paper to check the pH). Dilute an aliquot of the neutralized NTP appropriately, and read the optical density at the wavelengths given in the table below. Calculate the actual concentration of each NTP. Dilute the solutions with H2O to a final concentration of 50 mM NTP. Store each separately at -70°C in small aliquots.
| Base | Wavelength (nm) | Extinction Coefficient (E)(M-1cm-1) | ||
| A | 259 | 1.54 x 104 | ||
| G | 253 | 1.37 x 104 | ||
| C | 271 | 9.10 x 103 | ||
| T | 267 | 9.60 x 103 |
For a cuvette with a patch length of 1cm, absorbance = EM.
