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Application of reverse dot blot hybridization to simultaneous detection and identification of harmful algae
Chen, Guo Fu1,2; Zhang, Chun Yun1,2; Wang, Yuan Yuan1; Chen, Wen1
2015-07-01
Source PublicationENVIRONMENTAL SCIENCE AND POLLUTION RESEARCH
Volume22Issue:14Pages:10516-10528
SubtypeArticle
AbstractWarning and monitoring projects of harmful algal blooms require simple and rapid methods for simultaneous and accurate detection and identification of causative algae present in the environmental samples. Here, reverse dot blot hybridization (RDBH) was employed to simultaneously detect several harmful algae by using five representative bloom-forming microalgae along the Chinese coast. A set of specific probes for RDBH were developed by PCR, cloning, and sequencing of the internal transcribed spacer (ITS), alignment analysis, and probe design. Each probe was oligo (dT)-tailed and spotted onto positively charged nylon membrane to make up a low-density oligonucleotide array. Universal primers designed within the conserved regions were used to amplify the ITS sequences by using genomic DNA of target as templates. The digoxigenin (Dig)-labeled PCR products were denatured and then hybridized to the oligonucleotide array. The array produced a unique hybridization pattern for each target species differentiating them from each other. The preparations of oligonucleotide array and hybridization conditions were optimized. The developed RDBH demonstrated a detection limit up to 10 cells. The detection performance of RDBH was relatively stable and not affected by non-target species and the fixation time of target species over at least 30 days. The RDBH could recover all the target species from the simulated field samples and target species confirmed by the subsequent microscopy examination in the environmental samples. These results indicate that RDBH can be a new technical platform for parallel discrimination of harmful algae and is promising for environmental monitoring of these microorganisms.
KeywordHarmful Algal Blooms Internal Transcribed Spacer Reverse Dot Blot Hybridization Detection
DOI10.1007/s11356-015-4141-3
Indexed BySCI
Language英语
WOS IDWOS:000357344300014
Citation statistics
Document Type期刊论文
Identifierhttp://ir.qdio.ac.cn/handle/337002/48983
Collection海洋生态与环境科学重点实验室
Affiliation1.Harbin Inst Technol, Sch Marine Sci & Technol, Weihai, Shandong, Peoples R China
2.Chinese Acad Sci, Inst Oceanol, Key Lab Marine Ecol & Environm Sci, Qingdao 266071, Peoples R China
First Author AffilicationInstitute of Oceanology, Chinese Academy of Sciences
Recommended Citation
GB/T 7714
Chen, Guo Fu,Zhang, Chun Yun,Wang, Yuan Yuan,et al. Application of reverse dot blot hybridization to simultaneous detection and identification of harmful algae[J]. ENVIRONMENTAL SCIENCE AND POLLUTION RESEARCH,2015,22(14):10516-10528.
APA Chen, Guo Fu,Zhang, Chun Yun,Wang, Yuan Yuan,&Chen, Wen.(2015).Application of reverse dot blot hybridization to simultaneous detection and identification of harmful algae.ENVIRONMENTAL SCIENCE AND POLLUTION RESEARCH,22(14),10516-10528.
MLA Chen, Guo Fu,et al."Application of reverse dot blot hybridization to simultaneous detection and identification of harmful algae".ENVIRONMENTAL SCIENCE AND POLLUTION RESEARCH 22.14(2015):10516-10528.
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