qPCR

[xiaolong88]

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? 🙄

Hope you could help me 🙂

[JackBean]

you should first make the cDNA as soon after RNA extraction as possible and then you can handle it as you want 😉

[bravebeaker]

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

[Author]

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