Development of blocking oligos for avoiding non-target amplification in amplicon sequencing

Investigations in many study systems are hindered by loss of essential sequencing depth due to amplification of non-target DNA, including the genes of hosts and non-target microorganisms. This issue requires urgent attention to address ecologically relevant problems using high throughput, high resolution microbial profiling. Here, we use a modular approach to amplicon sequencing to characterize 8 interchangeable loci from bacteria, fungi, oomycetes and other eukaryotic microorganisms in a single sequencing run. To block amplification of non-target DNA in the target loci, we introduce a simple, low cost and highly flexible method using standard oligonucleotides (blocking oligos).