Evaluation of five in situ lysis protocols for PCR amenable metagenomic DNA from mangrove soils

• Five different metagenomic DNA extraction methods were compared on mangrove soils from three islands.
• Quantity and quality of the total community DNA were analysed.
• PCR amenability of isolated DNA was evaluated employing amplification of 16S rRNA gene.
• One among the protocols yielded PCR amenable DNA, while the protocol yielding highest concentration of DNA contained residual humic substance.

Microbes in nature are rarely amenable to growth by standard microbiological methods, with the majority being unculturable. Metagenomic methods help to bypass and overcome the limitations of traditional culturing method; wherein total community DNA is isolated, cloned into suitable vector and host systems. However, isolation of total community DNA itself remains a challenge. In this study five methods of total community DNA isolation from three different mangrove soils were evaluated to test its PCR amenability. The yield and purity of the isolated DNA was also analysed. The quantity of DNA by all 5 methods although reasonably high, contained residual humic contaminants. Of the five, the method employing liquid nitrogen yielded readily amplifiable DNA, while that by all others required further downstream processing to achieve purity and PCR amenability.