Truseq: insert size <100bp complications

achon · 10-14-2013, 06:59 PM

Hello community,
I currently have a designed custom amplicon panel with insert sizes around 100bp which for the most part work fine. I'm noticing some variability in that the longer insert sizes seem to have a consistently higher coverage than the shorter ones (lower vs upper quartile). Unfortunately qPCR and dPCR are not options due to the size of the panel.

Platform: MiSeq
Kit/Adapters: TruSeq

Before I redesign my primers, any advice? Is it possible to reliably reduce the insert size to ~50bp or lower and if so, what complications or design considerations are there? I've already lengthened the primer+adapters to somewhat already diminished fragment size.

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10-14-2013, 06:59 PM	#1
achon Junior Member Location: USA Join Date: Jun 2010 Posts: 1	Truseq: insert size <100bp complications Hello community, I currently have a designed custom amplicon panel with insert sizes around 100bp which for the most part work fine. I'm noticing some variability in that the longer insert sizes seem to have a consistently higher coverage than the shorter ones (lower vs upper quartile). Unfortunately qPCR and dPCR are not options due to the size of the panel. Platform: MiSeq Kit/Adapters: TruSeq Before I redesign my primers, any advice? Is it possible to reliably reduce the insert size to ~50bp or lower and if so, what complications or design considerations are there? I've already lengthened the primer+adapters to somewhat already diminished fragment size.