
PCR condition:
This for 30 cycle....
Initial Denaturation 94 C 3 min
Denaturation 94 C 1 min
Annealing - 1 min
Extension 72 C 2 min
Final extension 72 C 10 min
Hold 16 C Forever
Annealing temperature:
Sample Annealing temp
2 - 46 C
3 - 48.2 C
4 - 49.8 C
5 - 51.6 C
6 - 53.5 C
7 - 55.5 C
8 - 57.2 C
9 - 58.8 C
result for this annealing temperate can refer to images... ( 1 indicate maker, 2 to 9 is PCR product according to annealing temperature)
Master Mix:
Reagent for 1 sample
ddH2O 15.8ul
10x PCR buffer 2.5ul
MgCI 25mMol 1.5ul
dNTP 10mMol 1ul
primer F1 10mol/umol 1ul
R1 10mol/umol 1ul
Taq polymerase 0.2ul
DNA template 2ul
Refer to the result, what is indicate A? This is good result or very complicate result…?What modification can I do for get better result( single band)…..?Thank for you help…!!

Edited by veniesh raj, 10 October 2009 - 12:42 AM.