Jumping the green wall: the use of PNA-DNA clamps to enhance microbiome sampling depth in wildlife microbiome research

As microbiome research moves away from model organisms to wildlife, new challenges for microbiome high throughput sequencing arise caused by the variety of wildlife diets. High levels of contamination are commonly observed emanating from the host (mitochondria) or diet (chloroplast). Such high contamination levels affect the overall sequencing depth of wildlife samples thus decreasing statistical power and leading to poor performance in downstream analysis. We developed an amplification protocol utilizing PNA-DNA clamps to maximize the use of resources and to increase the sampling depth of true microbiome sequences in samples with high levels of plastid contamination. We chose two study organisms, a bat (Leptonyteris yerbabuenae) and a bird (Mimus parvulus), both relying on heavy plant-based diets that sometimes lead to traces of plant-based faecal material producing high contamination signals from chloroplasts and mitochondria.
On average, our protocol yielded a 13-fold increase in bac...