Abstract
The world economy is currently moving towards more sustainable approaches. Lignocellulosic biomass has been widely used as a substitute for fossil sources since it is considered a low-cost bio-renewable resource due to its abundance and continuous production. Compost habitats presenting high content of lignocellulosic biomass are considered a promising source of robust lignocellulose-degrading enzymes. Recently, several novel biocatalysts from different environments have been identified using metagenomic techniques. A key point of the metagenomics studies is the extraction and purification of nucleic acids. Nevertheless, the isolation of high molecular weight DNA from soil-like samples, such as compost, with the required quality for metagenomic approaches remains technically challenging, mainly due to the complex composition of the samples and the presence of contaminants like humic substances. In this work, a rapid and cost-effective protocol for metagenomic DNA extraction from compost samples composed of lignocellulosic residues and containing high content of humic substances was developed. The metagenomic DNA was considered as representative of the global environment and presented high quality (> 99% of humic acids effectively removed) and sufficient quantity (10.5–13.8 µg g−1 of compost) for downstream applications, namely functional metagenomic studies. The protocol takes about 4 h of bench work, and it can be performed using standard molecular biology equipment and reagents available in the laboratory.
Graphical Abstract
Key points/Highlights
• Metagenomic DNA was successfully extracted from compost samples rich in humic acids
• The improved protocol was established by optimizing the cell lysis method and buffer
• Complete removal of humic acids was achieved through the use of activated charcoal
• The suitability of the DNA was proven by the construction of a metagenomic library
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The authors confirm that the datasets supporting the findings and conclusions of this study are available within the article and its supplementary information file.
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Acknowledgements
AC and JS acknowledge her research grants UMINHO/BPD/37/2018 and UMINHOBIM/2020/28, respectively, under the scope of the project LIGNOZYMES (POCI-01-0145-FEDER-029773). The authors also acknowledge the Portuguese composting units Terra Fértil and Lipor for kindly supplying the compost samples.
Funding
This study was supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UIDB/04469/2020 unit and the Project LIGNOZYMES—Metagenomics approach to unravel the potential of lignocellulosic residues towards the discovery of novel enzymes (POCI-01–0145-FEDER-029773).
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AC, AS and JS conducted the experiments. SS and EG conceived and designed research. JR, SS and LR analysed data. SS and LR obtained financial support and coordinated the research. AC wrote the manuscript. All authors read and approved the manuscript.
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Costa, Â.M.A., Santos, A.O., Sousa, J. et al. Improved method for the extraction of high-quality DNA from lignocellulosic compost samples for metagenomic studies. Appl Microbiol Biotechnol 105, 8881–8893 (2021). https://doi.org/10.1007/s00253-021-11647-7
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DOI: https://doi.org/10.1007/s00253-021-11647-7