Recombinase polymerase amplification combined with lateral flow dipstick for the rapid detection of Chattonella marina

被引:5
作者
Zhang, Chunyun [1 ]
Yang, Yuchen [1 ]
Liu, Fuguo [1 ,2 ]
Wang, Yuanyuan [1 ]
Chen, Guofu [1 ,2 ]
机构
[1] Harbin Inst Technol Weihai, Sch Marine Sci & Technol, Wenhua West Rd 2, Weihai 264209, Peoples R China
[2] Harbin Inst Technol, Sch Environm, Harbin 264209, Peoples R China
基金
中央高校基本科研业务费专项资金资助;
关键词
Chattonella marina; Internal transcribed spacer; Recombinase polymerase amplification; Lateral flow dipstick; Detection; HARMFUL ALGAL BLOOMS; ROLLING CIRCLE AMPLIFICATION; SENSITIVE DETECTION; REAL-TIME; RED TIDE; VIRUS; RAPHIDOPHYCEAE; ASSAY; QUANTIFICATION; IDENTIFICATION;
D O I
10.1007/s10811-022-02737-8
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
Chattonella marina (Raphidophyceae) is a common microalgal species that has frequently caused harmful algal blooms (HABs). Here, recombinase polymerase amplification (RPA) was combined with a lateral flow dipstick (RPA-LFD) to establish a rapid detection method for this alga, aimed at providing an effective early warning of C. marina-forming HABs. A pair of specific RPA primers and an LFD probe was designed to target the internal transcribed spacer (ITS) sequence of C. marina. The specificity test with 20 control microalgal species indicated that the RPA-LFD was highly specific. Sensitivity test showed that the established RPA-LFD was 10 - 100 times more sensitive than conventional PCR. The detection limit of RPA-LFD was as low as 9.5 x 10(-1) ng mu L-1 for genomic DNA, 1.17 x 10(3) copies mu L-1 for recombinant plasmid containing the inserted ITS sequence, and 10 cells mL(-1) for the crude DNA extract of C. marina. In addition, the RPA-LFD was proved to be more accurate and sensitive than conventional PCR in the detection of target cells from natural samples, exhibiting its ability to be applied in the actual in situ detection scenarios. In conclusion, the developed RPA-LFD is rapid, specific, sensitive, and promising for being used in the point-of-care testing.
引用
收藏
页码:1607 / 1620
页数:14
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