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Development and evaluation of a reverse dot blot assay for the simultaneous detection of common toxic microalgae along the Chinese coast
Zhang, Chun Yun1,2; Chen, Guo Fu1,2; Cai, Pan Pan1; Wang, Yuan Yuan1; Guo, Chang Lu1
2015-07-01
发表期刊HARMFUL ALGAE
卷号47期号:2015页码:86-96
文章类型Article
摘要Toxic microalgae currently pose a great threat to human health, ecosystem, fishery, tourism, and aquaculture along the Chinese coast. The detection of toxic microalgae by routinely monitoring natural waters is necessary to provide timely mitigation. Therefore, an effective, simultaneous detection protocol should be established for the simple, rapid, and accurate identification of causative algae. This study developed and evaluated a reverse dot blot assay (RDBA) combined with a low-density membrane-based DNA array for the rapid and simultaneous detection of toxic microalgae that are commonly distributed along the Chinese coast. The large subunit rDNA D1-D2 regions of the target species were first sequenced to design taxonomic probes. Probe specificity was validated by performing a cross-reactivity test with dot blot hybridization. The tailed probes were immobilized onto a nylon membrane to prepare a low-density DNA array for RDBA. The established detection procedure involved DNA extraction, biotin (Bio)-labeling of objective sequences by multiple polymerase chain reaction (M-PCR), RDBA, coloration, and judgment of hybridization by the naked eye. Bio-labeled primer-based labeling proved to be an economical and effective method to prepare Bio-labeled PCR products for RDBA. The detection limits of RDBA using the M-PCR-labeling products from DNA templates prepared by different methods were also compared, and a kit-based DNA extraction method displayed the lowest detection limit of 0.5 cells. Simulation results showed that RDBA can recover all target species and was not affected by background DNA. RDBA was proven effective, specific, and sensitive for the simultaneous detection of toxic microalgae in the field samples. Therefore, this method may be used in the field monitoring of natural samples. (C) 2015 Elsevier B.V. All rights reserved.
关键词Toxic Microalgae Detection Lsu Rdna Multiplex Pcr Reverse Dot Blot Assay
DOI10.1016/j.hal.2015.06.003
收录类别SCI
语种英语
WOS记录号WOS:000359027400010
引用统计
文献类型期刊论文
条目标识符http://ir.qdio.ac.cn/handle/337002/49790
专题海洋生态与环境科学重点实验室
作者单位1.Harbin Inst Technol Weihai, Coll Oceanol, Weihai 264209, Shandong, Peoples R China
2.Chinese Acad Sci, Inst Oceanol, Key Lab Marine Ecol & Environm Sci, Qingdao 266071, Peoples R China
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Zhang, Chun Yun,Chen, Guo Fu,Cai, Pan Pan,et al. Development and evaluation of a reverse dot blot assay for the simultaneous detection of common toxic microalgae along the Chinese coast[J]. HARMFUL ALGAE,2015,47(2015):86-96.
APA Zhang, Chun Yun,Chen, Guo Fu,Cai, Pan Pan,Wang, Yuan Yuan,&Guo, Chang Lu.(2015).Development and evaluation of a reverse dot blot assay for the simultaneous detection of common toxic microalgae along the Chinese coast.HARMFUL ALGAE,47(2015),86-96.
MLA Zhang, Chun Yun,et al."Development and evaluation of a reverse dot blot assay for the simultaneous detection of common toxic microalgae along the Chinese coast".HARMFUL ALGAE 47.2015(2015):86-96.
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