PicoGreen Assay

[TUCF JSS]

Hi Everyone,

I was wondering if (to save on money) anyone has found PicoGreen assays to have any meaningful correlation to cluster count. We perform bioanalysis on our samples to give a rough estimation, however qPCR has been most accurate. We were just hoping a cheaper method of validation was available.

Any thoughts or input would be helpful! Thanks!

[TonyBrooks]

We use Picogreen (QuBit) and a size estimate from the Bioanalyser to dilute for cluster generation. It generally works OK once you optimise how much to add to the flowcell. There is obviously greater variation in cluster density than if you used qPCR to quantify your library.
Also, some library prep protocols and applications are more variable than others, (MeDip-Seq samples especially).

[HESmith]

qPCR is most accurate, but we've gotten reproducible results using picogreen for quantification and Bioanalyzer for size determination (and detecting adapter dimer contamination). Library-to-library variation in cluster number is within the error limits of pipetting volumes, at least for RNA/DNA/ChIP-Seq samples (we have not yet tested the more baroque libraries).