Yeast Colony PCR
PROCEDURE
- Add 1 µl Zymolyase 100T 10 mg/ml to 9 µl H2O and suspend a yeast colony in this solution.
- Add 40 µl PCR reaction mix to the suspended colony:
5 µl 5 mM gene specific primer 1
5 µl 5 mM gene specific primer 2
5 µl 10 x Taq buffer
5 µl 2 mM dNTPs
0.5 µl 10 mg/ml BSA
1 µl Taq
18.5 µl H2O - PCR cycle profile:
94°C 4 min
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94°C 1 min
55°C 1 min
72°C 1 min per kb
>>>35 cycles
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72°C for 10 min - Load 10 µl or more on a gel.
MATERIALS AND SOLUTIONS
10 X Taq Buffer:
0.2 M TRIS pH 8.3
15 mM MgCl2
0.25 M KCl
0.5% Tween 20