Based on the conserved 16S rDNA sequence of Microcystis, Cyanobacteria and microcystin synthetase gene B(mcyB), three pairs of specific primer 209F/409R, 27F1/409R and MTR/MTF are selected. The primer 409R is shared in two pairs. A duplex PCR to detect Microcystis and Cyanobacteria and a triplex PCR to detect Microcystis, Cyanobacteria and mcyB are optimized. The threshold of primers for cell concentration is also studied to detect the sensitivity of these primers. Results showed that the optimal concentrations for duplex PCR and for triplex PCR are from 105 cells·mL-1 to 103 cells·mL-1, and the duplex PCR could be used directly to detect crude water samples from reservoirs. This study suggested that the multiplex PCR is a simple and practical approach, and could play an important role on monitoring of microcystin in water.
张占会1, 谢数涛1*, 韩博平1, 林少君1, 钟秀英2, 林桂花2. 全细胞多重PCR检测蓝藻、微囊藻及产毒微囊藻方法初探[J]. , 2005, 24(1): 31-34.
ZHANG Zhan-hui1, XIE Shu-tao1, HAN Bo-ping1, LIN Shao-jun1, ZHONG Xiu-ying2, LIN Gui-hua2. Primary studies on the detection of Microcystis, cyanobacteria and microcystin synthetase gene by the whole-cell multiplex PCR. , 2005, 24(1): 31-34.
[1] Lawton L A, Codd G A. 1991. Cyanobacteria (blue-green algae) toxins and their significance in UK and European waters [J]. J Inst Water Environ Manage, 5:461-465.
[2] Sivonen K, and Jones G. 1999. Toxic cyanobacteria in water: a guide to their public health consequences, monitoring, and management[A]. In: Chorus I and Bartram J (ed). Cyanobacterial toxins[C], London, United Kingdom: E & FN Spon. 41-111.
[7] Pan H, Song L R, Liu Y D, et al. 2002. Detection of hepatotoxic Microcystis strains by PCR with intact cells from both culture and environmental samples [J]. Arch Microbiol,78(6): 421-427
[8] Emina A. 2003. Evaluation of a new DNA amplification technique to screen cyanobacteria isolated from drinking water sources for microcystin toxins [D]. Norse Scientist,Northern Kentucky University: 39-44
[9] Neilan B A, Jacobs D, Del Dot T, et al. 1997. rRNA sequences and evolutionary relationships among toxic and nontoxic cyanobacteria of the genus Microcystis[J]. Int J Syst Bacteriol, 47: 693-697.
[11] Tillett D, Dittmann E, Erhard M, et al. 2000. Structural organization of microcystin biosynthesis in Microcystis aeruginosa PCC7806: an integrated peptide polyketide synthetase system[J]. Chem Biol, 7 (10): 753-764.
