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Pipetting Guidelines in qPCR | Troubleshooting Forum

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I read that you should mix reaction mixtures with the gDNA by pipetting up and down several times. Is this really necessary?

BioTechniques Molecular Biology Forum brings bench scientists together in an online community to ask for and share advice regarding laboratory problems. This week's highlighted question comes from the DNA and General PCR Methods forum.

Q: I'm starting a new project that deals with DNA copy numbers. I just read through a kit reference card, which says that I should mix the reaction mixture with the gDNA by pipetting up and down several times. I've done lots of qPCR, but this never occurred to me. Is this really necessary? Wouldn't everything get mixed when I centrifuge it?

A: For my qPCR assays, it is needed. I tested standard dilution series with and without mixing by pipetting up and down, and the quality...

A: I normally pipet up and down a few times and stir up the solution with the same pipet tip. The heating may help mixing the solution, but optimal amplification...

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