Has anyone tried using multiple staggered custom sequencing primers in order to increase diversity when amplicon sequencing? I know you can stagger primers in order to generate base diversity. Would using multiple staggered sequencing primers (ie. +1, 0, -1 base position shift) do the same trick or would it introduce a new set of problems? I'll be testing this on a miseq if that makes any difference.
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I've discussed this with coworkers and while it should be possible, it adds complexity in the setup/downstream analysis for relatively little benefit since you can also just add more phiX to offset your low diversity. You can get away with some incredibly low diversity libraries (i.e. all same sequence) using a 20% phiX spike-in.
Here's a paper that describes a similar strategy that is also cool!
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NGS has revolutionized clinical microbiology by offering new ways to identify and surveil pathogens without prior knowledge of the target. One popular approach is 16S rRNA and ITS amplicon sequencing, which targets specific microbial markers. But studies are increasingly incorporating shotgun metagenomics, a technique that sequences all DNA from a sample without targeting specific microbial markers. This approach allows more detailed taxonomic classification, detection...Today, 06:46 AM -
Tumors contain complex mixtures of cell populations that evolve in response to treatment and host-specific factors. Tissue biopsies provide only static snapshots of a tumor’s state. But what if we could follow that evolution over time using a minimally invasive technique?
Liquid biopsies can detect tumor-derived material circulating in the bloodstream, providing a way to study treatment response, detect emerging resistance, and discover new biomarkers. Circulating...10-14-2026, 09:55 AM
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