We have a theory that large libraries, specifically those with fragments trailing off to >2kb, do not quantitate accurately from Bioanalzyer. We think this because qPCR concentrations do not correlate. Has anyone else had this problem? It seems if we are able to remove the "tail", then qPCR matches a bit better with Bioanalzyer.
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Libraries over 2 kb don't tend to work on Illumina sequencers, so do you really care what the concentration of those molecules is?
Also, are you doing a TaqMan qPCR or SYBR? If it's SYBR, you have to factor in the average molecule size, so you're still dependent on the Bioanalyzer anyway.
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Originally posted by kgoglin View PostWe have a theory that large libraries, specifically those with fragments trailing off to >2kb, do not quantitate accurately from Bioanalzyer. We think this because qPCR concentrations do not correlate. Has anyone else had this problem? It seems if we are able to remove the "tail", then qPCR matches a bit better with Bioanalzyer.
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