scholarly journals Comparison of Five Detection and Quantification Methods for Phytophthora ramorum in Stream and Irrigation Water

Plant Disease ◽  
2016 ◽  
Vol 100 (6) ◽  
pp. 1202-1211 ◽  
Author(s):  
Lucy Rollins ◽  
Katie Coats ◽  
Marianne Elliott ◽  
Gary Chastagner
Keyword(s):  
Quantitative Polymerase Chain Reaction ◽  
Detection Threshold ◽  
Phytophthora Ramorum ◽  
Detection Methods ◽  
Regulatory Agencies ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Low Levels ◽  
Irrigation Runoff ◽  
Leaf Disks

Propagules of Phytophthora ramorum, the causal agent of sudden oak death (SOD) and ramorum blight, can be recovered from infested stream and nursery irrigation runoff using baiting and filtration methods. Five detection methods, including pear and rhododendron leaf baits, Bottle O’ Bait, filtration, and quantitative polymerase chain reaction (qPCR) performed on zoospores trapped on a filter were compared simultaneously in laboratory assays using lab or creek water spiked with known quantities of P. ramorum zoospores. The detection threshold for each method was determined and methods that could be used to quantify zoospore inoculum were identified. Filtration and qPCR were the most sensitive at detecting low levels of zoospores, followed by wounded rhododendron leaves, rhododendron leaf disks, and pear baits. Filtration, qPCR, and leaf disks were able to quantify P. ramorum zoospores ranging from 2 to 451 direct-plate CFU/liter while wounded leaves and pear baits appeared to be better at detection rather than quantification. The ability to detect and quantify P. ramorum inoculum in water will assist scientists, regulatory agencies, and nursery personnel in assessing the risk of spreading P. ramorum in nurseries and landscape sites where untreated infested water is used for irrigation.

scholarly journals Eliminating false positives in a qPCR assay for the detection of the uidA gene in Escherichia coli

2013 ◽  
Vol 11 (3) ◽  
pp. 382-386 ◽  
Author(s):  
Richard Kibbee ◽  
Natalie Linklater ◽  
Banu Örmeci
Keyword(s):  
Escherichia Coli ◽  
Quantitative Polymerase Chain Reaction ◽  
Qpcr Assay ◽  
Uida Gene ◽  
Chain Reaction ◽  
Taq Polymerase ◽  
E Coli ◽  
Gene Copies ◽  
Polymerase Chain ◽  
Low Levels

Due to contaminant Escherichia coli DNA present in recombinant Taq polymerase reagents, it is not possible to reliably detect low levels of E. coli in samples using the quantitative polymerase chain reaction (qPCR) assay. Native Taq polymerase was successfully used in this study to detect five uidA gene copies (5 fg of genomic DNA) of the uidA gene.

scholarly journals Comparison of culture-based, vital stain and PMA-qPCR methods for the quantitative detection of viable hookworm ova

2017 ◽  
Vol 75 (11) ◽  
pp. 2615-2621 ◽  
Author(s):  
P. Gyawali ◽  
J. P. S. Sidhu ◽  
W. Ahmed ◽  
P. Jagals ◽  
S. Toze
Keyword(s):  
Environmental Samples ◽  
Quantitative Measurement ◽  
Quantitative Polymerase Chain Reaction ◽  
Detection Methods ◽  
Quantitative Detection ◽  
Propidium Monoazide ◽  
Chain Reaction ◽  
Vital Stain ◽  
Polymerase Chain ◽  
Hookworm Ova

Accurate quantitative measurement of viable hookworm ova from environmental samples is the key to controlling hookworm re-infections in the endemic regions. In this study, the accuracy of three quantitative detection methods [culture-based, vital stain and propidium monoazide-quantitative polymerase chain reaction (PMA-qPCR)] was evaluated by enumerating 1,000 ± 50 Ancylostoma caninum ova in the laboratory. The culture-based method was able to quantify an average of 397 ± 59 viable hookworm ova. Similarly, vital stain and PMA-qPCR methods quantified 644 ± 87 and 587 ± 91 viable ova, respectively. The numbers of viable ova estimated by the culture-based method were significantly (P < 0.05) lower than vital stain and PMA-qPCR methods. Therefore, both PMA-qPCR and vital stain methods appear to be suitable for the quantitative detection of viable hookworm ova. However, PMA-qPCR would be preferable over the vital stain method in scenarios where ova speciation is needed.

scholarly journals Event-Specific Qualitative and Quantitative Polymerase Chain Reaction Methods for Detection of Genetically Modified Rapeseed Ms8Rf3 Based on the Right Border Junctions

2008 ◽  
Vol 91 (1) ◽  
pp. 143-151 ◽  
Author(s):  
Gang Wu ◽  
Yuhua Wu ◽  
Ling Xiao ◽  
Changming Lu
Keyword(s):  
Polymerase Chain Reaction ◽  
Genetically Modified ◽  
Quantitative Polymerase Chain Reaction ◽  
Artificial Chromosome ◽  
Absolute Quantification ◽  
Detection Methods ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
The Absolute ◽  
The Right

Abstract Ms8Rf3 is a genetically modified rapeseed hybrid which is widely cultivated in Canada and exported to some other countries for production of foodstuffs or fodder. In this study, the genomic sequences flanking the right borders of the integrated transgenic sequences in the Ms8Rf3 genome were characterized and showed high similarities with the bacterial artificial chromosome clone of Chinese cabbage. Event-specific qualitative polymerase chain reaction (PCR) methods were established with the primers and probes targeting the junction regions to produce a 123 base pair (bp) product for the Ms8 event and 92 bp for the Rf3 event. The absolute detection limit of qualitative PCR was 2.5 initial template copies for the Ms8 event and 50 copies for the Rf3 event. Quantitiative detection methods were established, with the absolute quantification limit being approximately 25 initial template copies.

scholarly journals Polymerase chain reaction detection methods of Survival Motor Neuron genes: a review

2020 ◽  
Vol 1 (1) ◽  
pp. 37-43
Author(s):  
Azra Alimanović ◽  
Jasmin Šutković
Keyword(s):  
Polymerase Chain Reaction ◽  
Spinal Muscular Atrophy ◽  
Motor Neuron ◽  
Muscular Atrophy ◽  
Quantitative Polymerase Chain Reaction ◽  
Survival Motor Neuron ◽  
Detection Methods ◽  
Chain Reaction ◽  
Pcr Rflp ◽  
Polymerase Chain

Two SMN (survival motor neuron) genes are presented in the human genome: SMN1,  which present the telomeric gene whose homozygous deletion or mutation like gene conversion, causes spinal muscular atrophy (SMA), and SMN2, the centromeric version whose copy number modulates the phenotype of SMA These genes are commonly detected by Polymerase Chain reaction-based methods, and these are MLPA (Multiplex ligation-dependent probe amplification), qPCR (quantitative Polymerase chain reaction) and PCR-RFLP (Polymerase chain reaction-Restriction fragment length polymorphism). This paper reviews the current standing of the most common PCR methods used in the detection of spinal muscular atrophy genes. MLPA, qPCR, and PCR-RFLP currently represent the most common methods of choice for the detection of mutations, especially for deletion and duplication mutations.

scholarly journals Viability determination of Ascaris ova in raw wastewater: a comparative evaluation of culture-based, BacLight Live/Dead staining and PMA-qPCR methods

2019 ◽  
Vol 80 (5) ◽  
pp. 817-826 ◽  
Author(s):  
Vivek B. Ravindran ◽  
Esmaeil Shahsavari ◽  
Sarvesh K. Soni ◽  
Andrew S. Ball
Keyword(s):  
Quantitative Polymerase Chain Reaction ◽  
Comparative Assessment ◽  
Detection Methods ◽  
Propidium Monoazide ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Potential Public Health ◽  
Viability Determination ◽  
Raw Wastewater

Abstract Accurate evaluation of viable Ascaris ova in wastewater is the key to mitigating Ascaris reinfections in endemic regions. In this study, the viability of Ascaris ova in raw wastewater was determined using three different detection methods: culture-based, BacLight Live/Dead staining and propidium monoazide–quantitative polymerase chain reaction (PMA-qPCR). Furthermore, comparative assessment of viability utilising the aforementioned detection methods was performed using seeded experiments in wastewater. The percentage of viability was: culture-based (82%), BacLight Live/Dead staining (87%) and PMA-qPCR (85%) respectively. Despite the fact that no statistical difference was shown in the viability determination among the three methods, PMA-qPCR-based viability determination would be preferable over the other two methods for evaluating potential public health risks with A. suum ova due to its accuracy, being least subjective and its rapid reaction time.

Effect of zinc pyrithione shampoo treatment on skin commensal Malassezia

2020 ◽  
Author(s):  
Cheryl Leong ◽  
Joyce Wang ◽  
Min Jet Toi ◽  
Yuen In Lam ◽  
Joleen P Z Goh ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Quantitative Polymerase Chain Reaction ◽  
Detection Methods ◽  
Chain Reaction ◽  
Zinc Pyrithione ◽  
Qpcr Analysis ◽  
Malassezia Globosa ◽  
Polymerase Chain ◽  
Antifungal Efficacy ◽  
Malassezia Restricta

Abstract Malassezia restricta and Malassezia globosa are lipid dependent commensal yeasts associated with dandruff. Antifungal actives such as zinc pyrithione are commonly used in antidandruff shampoos, although their efficacy is not clearly demonstrated. In this study, we assessed the efficacy of antifungal treatments on scalp Malassezia via a combination of culturomic and genomic detection methods. Zinc pyrithione inhibited Malassezia growth at low minimum inhibitory concentrations (MICs). In a longitudinal pilot study, quantitative polymerase chain reaction (qPCR) analysis showed a decrease in M. restricta on the scalp after zinc pyrithione treatment. These findings validate the antifungal efficacy of zinc pyrithione as a dandruff treatment.

AN IMPROVED METHOD FOR INHIBITOR FREE NUCLEIC ACIDS FROM POLYPHENOL RICH LEAVES OF MUSA SPP

2017 ◽  
Vol 23 (1) ◽  
Author(s):  
N.NANDHA KUMAR ◽  
K. SOURIANATHA SUNDARAM ◽  
D. SUDHAKAR ◽  
K.K. KUMAR
Keyword(s):  
Polymerase Chain Reaction ◽  
Quantitative Polymerase Chain Reaction ◽  
Proteinase K ◽  
Chain Reaction ◽  
Banana Plant ◽  
High Molecular Weight Dna ◽  
Musa Spp ◽  
Leaf Tissues ◽  
Polymerase Chain

Excessive presence of polysaccharides, polyphenol and secondary metabolites in banana plant affects the quality of DNA and it leads to difficult in isolating good quality of DNA. An optimized modified CTAB protocol for the isolation of high quality and quantity of DNA obtained from banana leaf tissues has been developed. In this protocol a slight increased salt (NaCl) concentration (2.0M) was used in the extraction buffer. Polyvinylpyrrolidone (PVP) and Octanol were used for the removal of polyphenols and polymerase chain reaction (PCR) inhibitors. Proteins like various enzymes were degraded by Proteinase K and removed by centrifugation from plant extract during the isolation process resulting in pure genomic DNA, ready to use in downstream applications including PCR, quantitative polymerase chain reaction (qPCR), ligation, restriction and sequencing. This protocol yielded a high molecular weight DNA isolated from polyphenols rich leaves of Musa spp which was free from contamination and colour. The average yields of total DNA from leaf ranged from 917.4 to 1860.9 ng/ìL. This modified CTAB protocol reported here is less time consuming 4-5h, reproducible and can be used for a broad spectrum of plant species which have polyphenol and polysaccharide compounds.

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