Abstract
1. Nucleic acid extraction from complex environmental and ancient tissue material is prone to co-extract inhibitory substances that make further molecular analysis difficult or impossible. This co-extraction occurs in both solid-phase and liquid-phase/organic nucleic acid extraction protocols. Currently, the widely used method to overcome inhibition includes the addition of bovine serum albumin (BSA) to the downstream enzymatic reactions or the dilution of the nucleic acid extracts. BSA, however, seems to reduce the inhibitory effect of certain compounds only, and excessive dilution may change the original DNA composition.
2. In this study, we introduce an innovative new method using linear polyacrylamide (LPA) to efficiently precipitate and purify nucleic acids extracted from complex environmental and ancient tissue samples in one working step. The LPA method replaces the precipitation step in classic liquid-phase/organic extraction protocols or can be easily applied as an additional post-extraction step on impure DNA extracts. As a proof of concept, we experimented with this method on different ancient human mummy samples (bones, soft tissues and gut contents) from different time periods (5000 BC–1800 AD), as well as on complex environmental samples (e.g. soil, activated sludge and animal faeces) known to contain inhibitory compounds.
3. We demonstrated that LPA precipitates nucleic acids, even in an aqueous ethanol solution without the addition of chaotropic salts, resulting in the recovery of highly pure DNA from all tested samples that displayed inhibition with previously published extraction protocols. Compared to the current, most widely
used silica-based extraction method for ancient and sedimentary DNA, our LPA method resulted in comparable DNA qualities and overall DNA compositions (human endogenous content and microbial diversity).
4. In conclusion, our LPA method with its high purifying capacity provides an important alternative to the commonly used DNA extraction protocols in the environmental and ancient DNA (aDNA) fields.