DNA extraction from benthic Cyanobacteria: comparative assessment and optimization

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dc.contributor.authorGaget, V.
dc.contributor.authorKeulen, A.
dc.contributor.authorLau, M.
dc.contributor.authorMonis, P.
dc.contributor.authorBrookes, J.
dc.date.issued2017
dc.description.abstractAims: Benthic Cyanobacteria produce toxic and odorous compounds similar to their planktonic counterparts, challenging the quality of drinking water supplies. The biofilm that benthic algae and other micro-organisms produce is a complex and protective matrix. Monitoring to determine the abundance and identification of Cyanobacteria, therefore, relies on molecular techniques, with the choice of DNA isolation technique critical. This study investigated which DNA extraction method is optimal for DNA recovery in order to guarantee the best DNA yield for PCR-based analysis of benthic Cyanobacteria. Methods and Results: The conventional phenol–chloroform extraction method was compared with five commercial kits, with the addition of chemical and physical cell-lysis steps also trialled. The efficacy of the various methods was evaluated by measuring the quantity and quality of DNA by UV spectrophotometry and by quantitative PCR (qPCR) using Cyanobacteriaspecific primers. The yield and quality of DNA retrieved with the commercial kits was significantly higher than that of DNA obtained with the phenol– chloroform protocol. Conclusions: Kits including a physical cell-lysis step, such as the MO BIO Power Soil and Biofilm kits, were the most efficient for DNA isolation from benthic Cyanobacteria. Significance and Impact of the Study: These commercial kits allow greater recovery and the elimination of dangerous chemicals for DNA extraction, making them the method of choice for the isolation of DNA from benthic mats. They also facilitate the extraction of DNA from benthic Cyanobacteria, which can help to improve the characterization of Cyanobacteria in environmental studies using qPCRs or population composition analysis using next-generation sequencing.
dc.description.statementofresponsibilityV. Gaget, A. Keulen, M. Lau, P. Monis, and J.D. Brookes
dc.identifier.citationJournal of Applied Microbiology, 2017; 122(1):294-304
dc.identifier.doi10.1111/jam.13332
dc.identifier.issn1364-5072
dc.identifier.issn1365-2672
dc.identifier.orcidGaget, V. [0000-0001-5331-8600]
dc.identifier.orcidBrookes, J. [0000-0001-8408-9142]
dc.identifier.urihttp://hdl.handle.net/2440/105156
dc.language.isoen
dc.publisherWiley
dc.relation.granthttp://purl.org/au-research/grants/arc/LP120200587
dc.rights© 2016 The Society for Applied Microbiology
dc.source.urihttps://doi.org/10.1111/jam.13332
dc.subjectbenthic Cyanobacteria; DNA extraction; epilithic biofilm; MO BIO; phenol–chloroform; Qiagen
dc.titleDNA extraction from benthic Cyanobacteria: comparative assessment and optimization
dc.typeJournal article
pubs.publication-statusPublished

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