Evaluation of different DNA sampling techniques for the application of the real-time PCR method for the quantification of cyanobacteria in water.
OBJECTIVE
To evaluate different types of sample storage and DNA extraction techniques for the real-time PCR quantification of cyanobacteria in water.


RESULTS
Two different filter types for the cell harvest of Microcystis sp. and Planktothrix spp. that were either freeze-dried or stored frozen, and two different methods for DNA extraction were compared. DNA extraction was achieved by standard phenol-chloroform extraction or by a faster commercially available purification kit (DNeasy, QIAGEN). In general there was good agreement between the cell number equivalents of phycocyanin (PC) genotypes that were estimated using the Taq nuclease assay (TNA) between both filter types and the storing of samples. The standard DNA extraction procedure gave higher numbers of PC genotypes when compared with the DNeasy procedure. TNA results obtained from Planktothrix from natural samples extracted with the standard procedure revealed a significant correlation with the cell numbers estimated via the microscope.


CONCLUSIONS
Freeze-drying of samples gives quantifiable data. The standard DNA extraction is considered to be the most reliable and accurate, although the DNeasy procedure is useful for early warning monitoring.


CONCLUSIONS
Application of quantitative genotype analysis in cyanobacteria from freeze-dried samples collected during recent and past sampling programmes.
Schober
E
E
Institute for Limnology, Austrian Academy of Sciences, Mondsee, Austria.
Kurmayer
R
R
D015169:Colony Count, Microbial / Q000379:methods*; D000458:Cyanobacteria / Q000302:isolation & purification*; D004247:DNA / Q000302:isolation & purification*; D005374:Filtration; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013048:Specimen Handling; D019914:Taq Polymerase; D014871:Water Microbiology
D004247:DNA; D019914:Taq Polymerase