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Old 04-22-2015, 06:41 AM   #2
microgirl123
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Location: New England

Join Date: Jun 2012
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I've had several investigators do this with 18s. The Illumina/Nextera method works with any amplicon. You just need to be sure to pick locus primers that make an amplicon of an appropriate size. Illumina claims that you only need a 50 bp overlap - our experience with the 600-cycle kit is that the quality tanks at the ends of the reads so you want a much greater overlap.

The hardest part of the Illumina/Nextera method is getting your first round of PCR to work, especially with environmental samples. We generally recommend ordering just the locus-specific primers (without the partial adapters) first because they're cheap, optimizing your PCR with them, and then ordering the expensive tailed adapters.
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