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Old 08-10-2019, 08:07 AM   #9
Rosmano
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Location: Portugal

Join Date: May 2012
Posts: 18
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Those two bands with less than 100bps are adapter dimers.

I am having problems in the step of pre-amplification.
After pre-amplification I get a DNA quantity of 7.6ng/uL from 50,000 total thymocytes. However, when I try to run a qPCR I get no amplification in the 20 cycles. I ordered the TDE1 individually and directly from Illumina now that they discontinued the Nextera DNA kit, so I wasn't expecting to have problems with it.
The primers I am using are the ones from the Buenruostro paper. I'm at a loss for why I am not getting amplification in the qPCR to determine how many extra cycles I need. It seems my problem is with the primers.

What's the usual DNA concentration one has for 50,000 cells after TN5 treatment and after 5-cycle pre-amplification?
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