DNTP Recipe

From FreeBio

{Note: The following recipe makes fifty 50ul aliquots of 10mM dNTP. Deoxyribose triphosphates (dNTPs) differ from the normal DNA nucleotides in having three phosphates, instead of one. The breaking of the bond between the first and second phosphate group generates the energy needed to attach a new nucleotide to the polymer.}

1. In a 1.5ml (Eppendorf) tube, mix the following to get 2.5x dNTP stock:
   • 250ul 100mM dATP
   • 250ul 100mM dTTP
   • 250ul 100mM dGTP
   • 250ul 100mM dCTP.
2. Aliquot 500ul into two 1.5ml tubes.
3. Add 750ul ddH₂O to each of the two tubes to dilute to 10mM.
4. Aliquot 10mM dNTP into 50 tubes, 50ul per tube.
   {Note: Small aliquots are made in order to reduce the number of times that dNTP mixes are thawed and frozen, which degrade the nucleotides.}
5. Store @ -20C.