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Old 11-22-2019, 08:47 AM   #2
kmcarr
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Location: USA, Midwest

Join Date: May 2008
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Quote:
Originally Posted by ECorrales View Post
Hi! I have 24 ATAC libraries prepared for sequencing. For this, I want to pool them all together. I ran them on the TapeStation and I have the distinctive nucleosomal banding pattern (see attached files). I'm hesitating a bit on how to properly estimate the molarity of every library when I have multiple bands, since the Tape is not always detecting the same bands. Any suggestion about this? Should I just use Qubit quantification for pooling?
Use Region Analysis on the TapeStation software instead of Peak Analysis to determine an average size for each sample. Set up a Region from ~150-1000bp on all samples and use that calculate the average size. Use this average size and Qubit measurements for each to then calculate a molar concentration for each library.

Before you do this though, correct the Upper Marker in wells E1, A2, C2, F2 and G2 in the 10-24 file. The Upper Marker should be manually set tot he taller of the two peaks (Upper Marker peak height should alway be almost 2X the Lower Marker peak height.)
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