Math is not my strong point! I've pooled 12 samples, previously quantified by KAPA quant, based on their size-adjusted concentration. My library peaks average 700 bp. The KAPA quant of the pooled, size-adjusted library is less than I expected (not super low, just a bit lower than I was aiming for). Was the problem just pipetting or should I have pooled based on the non-adjusted concentration? I'm not sure that I haven't adjusted for size twice, and I don't want to overload the MiSeq.
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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,...10-30-2026, 06:46 AM