siRNA gone bad?

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GFPDude
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siRNA gone bad?

Hey everyone...
I have been using reverse transfection with invitrogen siRNA and Hiperfect transfection reagent to knockdown SRC-3 and SMRT in HeLa cells.

I have gotten the knockdown to work twice before, but then the knockdowns no longer work. Is the siRNA going bad? When its not in the freezer its on ice, but I do not know what is happening to the knockdown. It goes from 90% knockdown two experiments in a row to no knockdown from the 3rd experiment on.

Has anyone else had any similar problems?

Ivan Delgado
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From the way you describe your issue it sounds to me like your siRNA is going cycles of freezing and thawing. If that is the case, likely your siRNA is degraded. I strongly recommend splitting your siRNA into aliquots in such a way that you never have to re-freeze it more than once after it is used (thawed). 
 

Jason King
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Did you freeze the

Did you freeze the transfection reagent too? Most of these are stored at 4 degrees - I'm not familiar with HiPerFect though.

GFPDude
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Thanks Ivan. It may help to

Thanks Ivan. It may help to aliquot it before hand.
 
The Hiperfect reagent is just stored at 4 degrees. The siRNA is frozen.