Luckau Protocols:dNTPs

dNTPs

Tara K. Luckau's Home Page Conservation Genetics Lab Notebook Tara's Protocols

Purpose

dNTPs are a mixture of each of the nucleic acids used as building blocks of DNA strand synthesis during PCR. The mixture contains equal amounts of adenosine, guanine, cytosine and thymine such that each base is at a concentration of 2.5mM (10mM total) for direct use in PCR reactions.

Materials

Invitrogen 100mM dNTP Set

• catalog# 10297-018
• given: 250µL of 25µmol in H₂O @ 100mM each
• Media:Invitrogen 100mMdNTPProductManual.pdf

Protocol

Calculations

• Want: 10mM dNTP mix (2.5mM each), 5 aliquots of 1000µL
• (C₁) (V₁) = (C₂) (V₂)

(2.5mM) (5000µL) = (100mM) (V₂)

V₂ = 125µL of each dNTP

Make dNTP Mix

• 125µL dATP + 125µL dTTP + 125µL dCTP + 125µL dGTP + 4500µL H₂O = 5000µL dNTP Mix @ 10mM (2.5mM each dNTP)
• Aliquots: 1000µL into each of 5 snap-caps

4 stored in freezer

1 stored in fridge for immediate use