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Old 05-01-2019, 03:25 PM   #5
Junior Member
Location: Melbourne, Australia

Join Date: Aug 2013
Posts: 8

I haven't been checking the pH. It seems like the best place to start.
I do use calibrated pipettes, but I also have an addition dilution step that reduces the final concentration of NaOH to 0.4 mM. Does anyone have any thoughts on my process?

For denaturing and dilution:
- start with a 10 nM library and 0.2N NaOH (library: 10,000 pM, NaOH: 200 mM)
- 1:1 dilution with 0.2N NaOH (library: 5000 pM, NaOH: 100 mM)
- Add 990 uL HT1 (library: 50 pM, NaOH: 1 mM)
- Dilute library to 20 pM = 240 uL library:360 uL HT1 (library: 20 pM, NaOH: 0.4 mM)
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