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RT-qPCR Experimental Design


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#1 drabubakar

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Posted 03 December 2010 - 08:21 AM

Hello everyone,

I want to perform RT-qPCR.
I have silenced one gene (gene A) with four different siRNAs. I performed triplicate experiment for each siRNA.
I used GAPDH as a positive control to check for siRNA delivery efficiency. I performed triplicate experiment for GAPDH too.
I used scrambled siRNA as a negative control. I performed triplicate experiment for scrambled siRNA too.
Please I need help on the way I should arrange the RT-qPCR for:

1. Each 4 samples of the 4 siRNAs = 12 samples
2. The 3 samples of the GAPDH siRNA = 3 samples
3. the 3 samples of the scrambled siRNA = 3 samples
4. Non-template control (NTC) = ? samples

I was informed that I should use another house keeping gene such as ACTB that was not silenced to serve as a control for the Gene A and GAPDH which I have silenced.
How do I peeform QC for the transfection reagents I used?
I would be happy to receive help from everyone.

Thank you very much,

Dr. Abubakar

#2 drabubakar

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Posted 03 December 2010 - 08:23 AM

Please correction

1. Each 4 samples of the 4 siRNAs = 12 samples should be 1. Each 3 samples of the 4 siRNAs = 12 samples

Thank you,

Dr. Abubakar




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