I have had a problem with base calling due to a low diversity library on Miseq. I used P5 and P7 adapters with two amplification steps. The library is a PCR product. Would Y adapters be a better option, or am I not on the right track.
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Originally posted by nrobic View PostI have had a problem with base calling due to a low diversity library on Miseq. I used P5 and P7 adapters with two amplification steps. The library is a PCR product. Would Y adapters be a better option, or am I not on the right track.
1- Sequencing pool of different amplicons
2- Spiking in a high diversity library
3- increasing amplicons library diversity by adding 0-3 diversity nucleotides to 5' end of target specific sequences of primers (3' to adapter overhang)
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My first experiment with Y-adapters shows the base caller problem to be minimal as evaluated by the read 1-2 base intensity graph in Illumina SAV. The variant caller also shows a great reduction in false variants. I still need to load my control for comparison and repeat this finding again - Thanks
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