Genetics Lab Procedures

(Lab 1) What are the basic components needed by PCR

1) DNA Template

2) Primers

3) Taq polymerase

4) Deoxyribonucleotide triphosphates (dNTPs)

5) Buffer solution

6) Divalent cations

7) Double-distilled H2O (ddH2O)

DNA Template

Starting material which will contain the target region of DNA that we wish to amplify

Primers

Needed to initiate DNA synthesis by the enzyme DNA polymerase, and which also determine the ends of the DNA fragments to be amplified

Taq polymerase

A DNA polymerase isolated from the bacteria Thermus aquaticus, which lives in hot springs (95-100°C). It’s used to synthesize many copies of the target DNA

Deoxyribonucleotide triphosphates (dNTPs)

DNA monomers that the polymerase uses to build new DNA polymers

Buffer solution

Maintains a stable and favorable salt concentration and pH for the Taq polymerase enzyme

Divalent cations

Often acts as enzyme cofactors and MgCl2 is required for the activity of strand extension by Taq polymerase (MgCl2 at 1-2.5mM)

Double-distilled H2O (ddH2O)

Extremely clean water that has been autoclaved, so it is both free of impurities and sterile, which is added to the PCR to bring all components to their final working concentrations