polymerase chain reaction

polymerase chain reaction (PCR)

1. A basic PCR set up requires several components and reagents.[8] These components include:

①DNA template that contains the DNA region (target) to be amplified.

②Two primers that are complementary to the 3' (three prime) ends of each of the sense and anti-sense strand of the DNA target.

Note: Primers can be sythesized by biotechnology (sequence synthesized) company after we submit our sequence of primers designed by us using software (Primer Primer 5.0) as long as we know the sequence of target DNA region.

③Taq polymerase or another DNA polymerase with a temperature optimum at around 70°C.

④Deoxynucleoside triphosphates (dNTPs, sometimes called "deoxynucleotide triphosphates"; nucleotides containing triphosphate groups), the building-blocks from which the DNA polymerase synthesizes a new DNA strand.

⑤Buffer solution, providing a suitable chemical environment for optimum activity and stability of the DNA polymerase.

⑥Bivalent cations, magnesium or manganese ions; generally Mg2+ is used, but Mn2+ can be utilized for PCR-mediated DNA mutagenesis, as higher Mn2+ concentration increases the error rate during DNA synthesis.

⑦Monovalent cation potassium ions.

Note: ③④⑤⑥⑦can be replaced by mix sold in biotechnology company (eg. Sigma or Genview)

2.Reagents & Instruments:

①PCR tubes : 0.2 ml, from Genview Company or others)

②DNA Marker: DL 2,000 or DL 15,000 (depended on the length of target DNA region)

③PCR-Cooler