LONG PCR

Reaction (100 µl total volume):

72 µl water

10 X PC2 Buffer - 10 µl

25 mM MgCl₂ - 4 µl

8 mM dNTPs - 10 µl

Primers (100 pmoles/µl each) - 2 µl

kDNA (100 ng/µl) - 1 µl

Klentaq (25 U/µl) - 0.5 µl

Pfu (2.5 U/µl) - 0.1 µl

Cycling conditions: 9600 Perkin Elmer Machine

25 cycles

Initial denaturation: 94oC/5 sec

Denaturation: 94oC/5 sec

Annealing: 50oC/20 sec

Elongation: 68oC/5 min

10 X PC2 Buffer (comes with Klentaq enzyme)

500 mM Tris HC1 (pH 9.1)

160 mM (NH4)2SO4

25 mM MgCl2

1.5 mg/ml BSA