Development of a chip assay and quantitative PCR for detecting microcystin synthetase E gene expression.
The chip and quantitative real-time PCR (qPCR) assays were optimized to study the expression of microcystin biosynthesis genes (mcy) with RNA samples extracted from cyanobacterial strains and environmental water samples. Both microcystin-producing Anabaena and Microcystis were identified in Lake Tuusulanjärvi samples. Microcystis transcribed the mcyE genes throughout the summer of 2006, while expression by Anabaena became evident later in August and September. Active mcyE gene expression was also detectable when microcystin concentrations were very low. Detection of Anabaena mcyE transcripts by qPCR, as well as certain cyanobacterial 16S rRNAs with the chip assay, showed slightly reduced sensitivity compared with the DNA analyses. In contrast, even groups undetectable or present in low quantities as determined by microscopy could be identified with the chip assay from DNA samples. The methods introduced add to the previously scarce repertoire of applications for mcy expression profiling in environmental samples and enable in situ studies of regulation of microcystin synthesis in response to environmental factors.
Sipari
Hanna
H
Department of Food and Environmental Sciences, Division of Microbiology, University of Helsinki, Viikki Biocenter 1, P.O. Box 56, FI-00014 Helsinki, Finland.
Rantala-Ylinen
Anne
A
Jokela
Jouni
J
Oksanen
Ilona
I
Sivonen
Kaarina
K
D017033:Anabaena / Q000201:enzymology* / Q000302:isolation & purification; D001426:Bacterial Proteins / Q000096:biosynthesis; D001431:Bacteriological Techniques / Q000379:methods; D004783:Environmental Microbiology; D020869:Gene Expression Profiling / Q000379:methods*; D008025:Ligases / Q000096:biosynthesis*; D052998:Microcystins / Q000096:biosynthesis*; D046931:Microcystis / Q000201:enzymology* / Q000302:isolation & purification; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
D001426:Bacterial Proteins; D052998:Microcystins; C078588:microcystin; D008025:Ligases
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