Rapid estimation of species-specific DNA digestibility based on differential qPCR

Digestibility of DNA in selected prey species was estimated by an in vivo feeding trial using six test diets. Each diet contained one of the following: (a) goldfish, (b) goldfish + citric acid, (c) goldfish + CaCO₃, (d) shrimp, (e) snail, and (f) goldfish + shrimp + snail. The diets were fed to rainbow trout and fecal samples were collected. Mitochondrial DNA was extracted from both diet and fecal samples and quantified by qPCR using two sets of species-specific primers for each prey species. The first set of primers amplified a short stretch of DNA (51–80 bp), whereas another set amplified a longer stretch (126–162 bp). DNA digestibility was estimated based on the ratio between short and long amplicons using the following formula: Digestibility (%) = log_{((Mt?S)/(Mt?L))} (S]/L])/(0.01 × Mt), where Mt: 16,000 (bp), S: length of short amplicon (bp), L: length of long amplicon (bp), S]: relative quantity of short amplicon, L]: relative quantity of long amplicon. Calculated fecal digestibility of goldfish DNA ranged from ?1.05 % (f) to 2.07 % (a). Digestibility of shrimp DNA was 10.79 % (d) and 12.61 % (f). Digestibility of snail DNA was 1.88 % (e) and 2.06 % (f). These results suggest that the digestibility of dietary DNA can be estimated based on the ratio between long and short fragments.