Hello Syed

You should use DEPC treated water. You need to avoid RNase contamination which may be present in autoclaved double distilled water. RNases get destroyed at temperatures greater than 180°C for more than 5-6 hours or by using chemicals like DEPC to treat water. Moreover, there may be other substances like minerals which could interfere with your qPCR reactions which require specific salt concentration. I would suggest you use one of these waters, DEPC treated water, nuclease-free water or RT-PCR grade water.

Thank you.

Hello Syed

You should use DEPC treated water. You need to avoid RNase contamination which may be present in autoclaved double distilled water. RNases get destroyed at temperatures greater than 180°C for more than 5-6 hours or by using chemicals like DEPC to treat water. Moreover, there may be other substances like minerals which could interfere with your qPCR reactions which require specific salt concentration. I would suggest you use one of these waters, DEPC treated water, nuclease-free water or RT-PCR grade water.

Thank you.

Yes you should use autoclaved water that has been DEPC treated first

Not just autoclaved water. Crucially autoclaving alone does NOT kill RNAses

To be on the safe side use nuclease-free water.

and you could run one sample using DEPC treated water and compare the results. Then share with us ur results, to see the effect.

thanks

Dear Malcolm Nobre , Laurence Stuart Dawkins-Hall and Hajar Alghamdi ,

Thanks a lot for your kind suggestions.

We normally autoclave DEPC treated water for subsequent applications. However, DEPC might inhibit PCR reactions and therefore, it would be better to use it only for RNA extraction and not PCR.

For PCR, you might use autoclaved millipore water or any other RNase free water.

Hi Syed,

DEPC has worked wonderfully for us for years now, including RNA isolation cDNA dilution and subsequent RT-qPCR. We typically have very clean melt curves.

Good luck!

I have no experience with DEPC treated water. But have used always autoclaved Mili q water and that has worked also pretty good. Good primer efficiency with cDNA and no negative contamination.

Dear Dr Sayanta Bera, thanks a lot for sharing your experience!

Normally, the protocol recommends DEPC, well I don't have experience using autoclaved water. During my experiments, I use only DPEC or grade water include in the RT-PCR kits.

Dear Naqash Goswami, thanks a lot for sharing your experience.

I have been using DEPC treated water for qPCR, and the result was OK.

Dear Zhongqing Li, Thank you for sharing your experience.

Dear Dr Sjon Hartman and Vajiheh Safavi Rizi, Thank you for sharing your experience.