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Old 11-30-2018, 12:51 PM   #1
kgoglin
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Location: San Diego

Join Date: Dec 2014
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Default TruSeq PCR-free libraries inefficient ligation of adapters

It seems to be well known that the ligation efficiency of the adapters for the TruSeq PCR-free library prep is low. Bioanalyzer concentrations can be 3-30 fold higher than qPCR. I believe the qPCR concentrations are accurate, however, the sequencing runs are under-clustered. I suspect the libraries are not getting denatured properly. Has anyone had this happen?

My theory is that the NaOH is being used up on the DNA fragments with no adapters and therefore less NaOH is available to denature the libraries.

Thanks,
Karrie
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