Had 32 primers made that started with "NNNN" at 5' end. I have a full lane of Illumina data reading through these bases and into an inline bar code. I almost always recover the bar code perfectly. I expected to see each base at ~25% in these cycles; instead I get 16%, 17%, 30% and 37% for A, C, G, and T, respectively. That's some pretty sloppy work by the company. I haven't made oligos myself in years, but isn't is just a matter of mixing the four bases in a 5th bottle "N" that goes on the synthesizer?
Interesting what you learn now that sequencing through oligos as single molecules common. Had a synthesis by a different company (one that is usually highly recommended) where the 3' base was incorrect. They are synthesized from 3' end.
Interesting what you learn now that sequencing through oligos as single molecules common. Had a synthesis by a different company (one that is usually highly recommended) where the 3' base was incorrect. They are synthesized from 3' end.
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