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Old 09-24-2012, 05:51 PM   #4
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Location: Monash University, Melbourne, Australia.

Join Date: Jan 2008
Posts: 246

I'm not saying that this is the solution to your problem, but a couple of things come to mind as possibilities:

I'd measure the amount of material in the PCR tube before and after cycling to make sure you're actually getting something produced from the PCR reaction and that the yield is good enough. Also measure the size distribution on a bioanalyzer after 'tagmentation' and/or PCR. Sometimes the size range with different samples is very large (i.e. libs with fragments down near 100b, others at 2kb) and this might affect the normalisation too; but, I haven't actually measured the effect of fragment size on normalisation...


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