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===Discussion=== Well, I'm not sure there is a clear indication, at least at the temperature range I used, that there is a temperature dependence. Comparing the 65C (theoretically the most stringent) and the 50C (least stringent) show little difference overall between the off-target bands. The lowered primer concentration does seem to help though, so I can try again on Monday with a continually decreasing level of primers. This also raises a point that the NTC consistently showed primer-dimer signal (~110bp). Looking back at some of the previous gels with NTCs ([[Daniel:Notebook/ComboLock/2016-8-19|this]] one shows it best), I think that the primer-dimers do form on their own but are helped by the presence of the padlock especially. Of note however is also that the number of cycles for this test was higher than usual. From previous PCR curves (see example from the [[Daniel:Notebook/ComboLock/2016-8-30|X3 dilution test]]) you can see that I went an extra 6 cycles or so and that the NTC lanes start to amplify around cycle 23. [[Category:ComboLock]] [[Category:20160922]]
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