水华蓝藻产毒特性的PCR检测法
CHARACTERIZATION OF TOXIC WATERBLOOM-FORMING CYANOBACTERIA BY MODIFIED PCR
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摘要: 特异引物对(TOX 1P/1F;TOX 2P/2F)用于检测微囊藻毒素合成酶基因mcyB片段在38种水华蓝藻中的分布情况.结果显示,所有能产生微囊藻毒素的微囊藻都有特异扩增条带,非产毒株则没有.几种常规的毒性检测方法验证了PCR方法所获结果的准确性.本研究发展了以全细胞PCR法检测mcyB片断,说明全细胞PCR检测法适用于不同来源的蓝藻材料.结果证明以DNA为基础鉴别产毒和非产毒微囊藻及其他水华蓝藻的方法是可行和实用的.Abstract: The present study surveyed the distribution of microcystin-producing gene-mcyB by PCR among 38 bloom-forming cyanobacterial strains from FACHB-Collection. The results demonstrated that special amplifications were being gotten only from 19 toxic Microcystis strains, whereas it was deficient in nontoxic ones got none. The PCR results showed highly agreement with the toxicity of every strain determined by HPLC, ELISA, and Bioassay respectively. Furthermore, the traditional PCR protocols were simplified by checking the cyanobacteria cells directly instead of the previously used extracted genomic DNA. The whole cells PCR we developed in this study could be successfully applied to cultured algae materials, water samples and lyophilized cyanobacterial cells. Thus, discrimination of toxic and nontoxic Microcystis strains by molecular biological technologies proves to be practicable and efficacious.
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Keywords:
- Whole cells PCR /
- Water-blooming cyanobacteria /
- Microcystis /
- Microcystins /
- mcyB /
- Gene
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the cyanobacterium Microcystis aeruginosa [J].Applied and Environmental Microbiology.1999,65(2):737—739[14] Neilan B A,Dittmann E,Borner T.Nonribosomal peptide synthesis and toxigenicity of cyanobacteria [J].J.Bacteriol.1999,181,13:4089—4097[15] Dittmann E,Neilan B A,Borner T.Peptide synthetase genes occur in various species of cyanobacteria [M].The Phototrophic Prokaryotes.New York:Kluwer Academic/Plenum Publishers 1999[16] Ichimura T.Media for blue-green algae [M].Methods in algalogical studies.Tokyo,Kyoritsu,1979[17] 董传辉,俞顺章,陈 刚.某湖周围水厂源水及出厂水微囊藻毒素调查[J].卫生研究.1998,27,3:100—102[18] 宋立荣,雷腊梅,何振荣.滇池水华蓝藻铜绿微囊藻和绿色微囊藻的生长生理特性及毒素分析[J].水生生物学报.1999,23(5):402—407[19] 何家菀,李络平,俞家禄,等.我国产毒微囊藻的新发现——韦氏微囊藻及其毒性的初步研究[J].水生生物学报.1996,20,2:192—194 Watanabe M F,Harada K,Carmichael W W,et al.Toxic Microcystis [M].New York:Boca Raton CRC Press 1996[2] Carmichael W W.The Toxins of Cyanobacteria [J].Scientific American.1994,274(1):78—86[3] Carmichael W W.Cyanobacterial Toxins [J].IOC Manuals and Guides.1996,33:163—175[4] Watanabe M F.,Oishi S.Toxicities of Microcystis aeruginosa collected from some lakes,reservoirs,ponds and moat in Tokyo and adjacent regions [J].Jpn.J.Limnol.1980,41:5—9[5] Shirai M K,Obotake M A.Toxicity and delayed type hypersensitivity caused by Microcystis blooms from lake Kasumigaura [J].Micrbiol Immunol.1986,30:731—735[6] Kato T,Watanabe M F,Watanabe M.Allozyme divergence in Microcystis(Cyanophyceae)and its taxonomic inference [J].Arch.Hyrobiol..Bd.1991,64(Suppl):129—140[7] Makoto S,Akio O,Takaki S,Seiishiro T.Toxicity and toxins of natural blooms and isolated strains of Microcystis sp.(Cyanobacteria)and improved procedure for purification of cultures [J].Applied and Environmental Microbiology.1991,4:1241—1245[8] Baker D.,Humpage A R.,Steffensen D A.Cyanobacterial blooms in the Murray-Basin:Their Taxonomy and Toxicity [M].Austr.Center for Water Quality Res.1993[9] Codd J N,Bettie K A,Lawton L A,et al.Blue-green algal toxins [J].British 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