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question about pcr


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3 replies to this topic

#1 iraj

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Posted 04 September 2010 - 10:58 AM

dear all

I working on goat and sheep promotor gene amplification.
I have a high quality DNA and I extracted it from blood.
One day I'll get the answer, but the next day with the same protocol does not get the answer in pcr
In the case I do many expriment but dont different in results.
please help me emergency

#2 cagdas

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Posted 05 September 2010 - 10:11 AM

Hi iraj,

You should check the following items.

1)Verify that you have not forgetten any reagents in your master mix.
2)PCR protocol?
3)Change dNTPs.
4)Check your primers for dimerization errors or hairpin formation.
5)DNA quality?

Did you use the same sample?

#3 iraj

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Posted 05 September 2010 - 05:51 PM

Hi iraj,

You should check the following items.

1)Verify that you have not forgetten any reagents in your master mix.
2)PCR protocol?
3)Change dNTPs.
4)Check your primers for dimerization errors or hairpin formation.
5)DNA quality?

Did you use the same sample?


thanks cagdas
but i checked all of them
result not changed!

#4 cagdas

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Posted 05 September 2010 - 11:48 PM

Hi iraj,

Did you change all the reagents?
How many samples do you analyze in one go?
What' s your pcr reaction condition, protocol and expected amplicon size?
Do you run your fragments out on acrylamide or agarose?
Is agarose at the correct % for the fragment sizes you are looking at?




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