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qPCR

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qPCR

Postby xiaolong88 » Tue Dec 11, 2012 4:58 pm

Hi,

I have transcript (RNA) which I would use for qRT PCR for standard curve-the standard.
I should quantitate the transcript (RNA) before reverse transcribe them into cDNA.
I wonder I should serially dilute the transcript and perform reverse transcription? Or I should synthesis the cDNA first and only perform serial dilution for qPCR? Any different? :roll:

Hope you could help me :)
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Postby JackBean » Tue Dec 11, 2012 11:00 pm

you should first make the cDNA as soon after RNA extraction as possible and then you can handle it as you want ;)
http://www.biolib.cz/en/main/

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Postby bravebeaker » Wed Dec 26, 2012 4:47 am

1. extracted RNA --> DNase treatment --> RT to synthesize cDNAs
2. use same primers to amplify your target by PCR
3. extract bands from gel and purify target DNA
4. use the purified DNA for making a standard curve.
5. runa a qPCR
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