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Old 10-31-2012, 12:24 PM   #7
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Location: California

Join Date: Jun 2012
Posts: 3

Pmiguels you are right, I took this off of the Nextera XT page at illumina....

Smaller amplicon inputs into Nextera XT preps typically yield smaller insert size ranges. To maximize recovery of smaller fragments out of the AMPureXP cleanup we recommend > 300 bp to ensure even coverage across the length of the DNA fragment. An expected drop off in sequencing coverage about 50 bp from each distal end of a fragment may be seen. This is because the tagmentation reaction cannot add an adapter right at the distal end of a fragment. For PCR amplicon sequencing this can be easily averted by simply designing your amplicons to be ~100 bases larger than the desired insert to be sequenced.

Are there any protocol differences based on different amplicon lengths?
For amplicons greater than 500bp, Illumina recommends using a 0.6x AMPure XP cleanup (30 μl volume of beads). For amplicons less than 500bp, Illumina recommends using a 1.8x AMPure XP cleanup (90 μl volume of beads) to maximize yield."
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