Simultaneous detection of harmful algae by multiple polymerase chain reaction coupled with reverse dot blot hybridization

Zhang, Chun Yun; Chen, Guo Fu; Liu, Yang; Wang, Yuan Yuan; Xu, Zhenbo; Zhang, Bao Yu; Wang, Guang Ce

doi:10.1016/j.hal.2014.03.004

Cited by 12 publications

(4 citation statements)

References 35 publications

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“…All probes for DBH were 5 0 -end-labeled with digoxigenin (Dig) in accordance with the methods described by Zhang et al (2014). Genomic DNA was extracted from algal cultures (Table 1) by using the Plant Genomic DNA Rapid Extraction Kit (Sangon Biotech, Shanghai, China) in accordance with the manufacturer's instructions.…”

Section: Experimental Verification Of Probe Specificitymentioning

confidence: 99%

Development and evaluation of a reverse dot blot assay for the simultaneous detection of common toxic microalgae along the Chinese coast

et al. 2015

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Section: Experimental Verification Of Probe Specificitymentioning

confidence: 99%

Development and evaluation of a reverse dot blot assay for the simultaneous detection of common toxic microalgae along the Chinese coast

et al. 2015

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“…Several DNA-based typing techniques, such as restriction fragment length polymorphism (Perrig et al, 2015;Tomazi et al, 2019), DNA random amplification reaction, pulsed field gel electrophoresis (PFGE; Reinoso et al, 2015;Fessia et al, 2018;Werner et al, 2018), and multilocus sequence typing (Coffey et al, 2006;Rato et al, 2008;Lang et al, 2009) have already been described for the characterization of S. uberis. However, methods based on specific probes derived by PCR (hybridization) provide higher accuracy for virus and bacterial discrimination and involve lower costs (Meng et al, 2007;Zhang et al, 2014;Guo et al, 2015). Unlike PCR, which uses target fragments or sequences for identification, DNA hybridization uses DNA denatured by heat, and depends on the positive hybridization signal of the target sample with known DNA probes.…”

Section: Introductionmentioning

confidence: 99%

Application of a dot blot hybridization assay for genotyping Streptococcus uberis from Brazilian dairy herds

Martins

Ribeiro

Tavares

et al. 2021

Journal of Dairy Science

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Streptococcus uberis is a major cause of environmental mastitis in many regions, and it is associated with clinical and subclinical infections. Although the main source of infection is the environment, reports of strains with a contagious profile have been described. Dot blot hybridization analysis allows the rapid identification of S. uberis population structures within and between herds, and it helps to identify strain diversity as well as possible clonal lineages that directly affect the control of bovine mastitis caused by this pathogen. The aim of this study was to evaluate the diversity of S. uberis isolates obtained from clinical (n = 22) and subclinical (n = 22) cases of mastitis in dairy herds (n = 13) in Brazil over a period of 12 mo. We submitted 44 S. uberis isolates to dot blot hybridization followed by automatic data analysis. We identified 8 different hybridization patterns using genetic markers associated with virulence factors and taxonomy, indicating diversity of S. uberis within the population and suggesting environmental transmission. However, the evidence of identical dot blot patterns in different mammary quarters from the same animal also suggested local contagious transmission. Of the virulence genes evaluated, we found a high prevalence of the genes sua, pauA, and gapC, highlighting the importance of these virulence factors for the adhesion, invasion, and multiplication of S. uberis in subclinical and clinical intramammary infections.

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“…Dot blot hybridization (DBH) is a method of nucleic acid molecular hybridization, which has been widely used in virus detection in recent years. [31][32][33] DBH technology refers to the hybridization of nucleic acid probe with nucleic acid DNA of the sample to be tested on the hybridization membrane. The nucleic acid probe is equipped with appropriate markers for post-reaction detection, and combines with specific target molecules to form a hybrid body, which is then displayed by color reaction.…”

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confidence: 99%

“…Affinity determination of aptamer to PFF.-Before preparation of the aptasensor, the affinity between Apt and PFF was measured by DBH referring to the previous literature of zhang et al 31 Using an Apt 3 as a probe template, 5 × Transcription Buffer, DTT (1 μM), DIG, T7 RNA polymerase and DEPC water were added into it. After mixing, they were put in water bath at 37 °C overnight, and 5 μl probe solution was obtained.…”

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A Dual-Amplification Electrochemical Aptasensor for Profenofos Detection

Zhang

Sun

Cheng

et al. 2020

J. Electrochem. Soc.

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The present study reported a dual-amplification electrochemical aptasensor for sensitive detection of profenofos (PFF) in vegetables. A screen-printed carbon electrode (SPCE) modified with graphitized multi-walled carbon nanotubes (MWCNTGr) and Au nanoshell was used as a test platform, which ensured a rapid detection process and showed a favorable electrochemical performance. MWCNTGr and Au nanoshell enhanced the electrical conductivity and the surface area, thus the detection signal was amplified. The affinity between PFF and its aptamer (Apt) was verified firstly by dot blot hybridization (DBH), and the result was exciting. Furthermore, the effects of the aptamers modified respectively with -NH2 and -SH on the current signal were compared with each other by cyclic voltammetry (CV), and results showed that the aptamers modified with -NH2 made the current signal change more obvious. Based on all above, a high-efficiency electrochemical aptasensor was fabricated with a wide linear range from 0.1–1 × 105 ng · ml−1 and a detection limit of 0.052 ng · ml−1 under the optimized conditions. This aptasensor had great specificity, stability and reproducibility. Hence, the developed aptasensor was successfully used to detect PFF in vegetables. The proposed method also has a potential for the detection of other organophosphorus pesticide (OPs).

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Simultaneous detection of harmful algae by multiple polymerase chain reaction coupled with reverse dot blot hybridization

Cited by 12 publications

References 35 publications

Development and evaluation of a reverse dot blot assay for the simultaneous detection of common toxic microalgae along the Chinese coast

Development and evaluation of a reverse dot blot assay for the simultaneous detection of common toxic microalgae along the Chinese coast

Application of a dot blot hybridization assay for genotyping Streptococcus uberis from Brazilian dairy herds

A Dual-Amplification Electrochemical Aptasensor for Profenofos Detection

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