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Old 11-18-2011, 06:55 AM   #19
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Location: Massachusetts

Join Date: May 2009
Posts: 116

Originally Posted by HESmith View Post
There's an alternative approach, assuming that you have not yet constructed the libraries. Design them so the junction is at the opposite end of the insert, and perform paired-end sequencing. Cluster calling is based only on the first five cycles of read one, so you'll avoid the low-complexity issue.

I have a sample of 96-plex low diversity amplicon libraries running now and clusters were found just fine--but the low diversity is causing a tremendous discrepancy between the blue and the green box-and-whiskers plot--raw clusters and clusters passing filter. I hope those data are recoverable at the end. Nothing in my primer design, barcoding, indexing scheme can change the fact that it's "low complexity". First four bases were completely random and followed by eight different in-line bar codes.

This is PE sequencing.

Yet I know labs are making this work.
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