scholarly journals Quantification of Agrobacterium vitis from Grapevine Nursery Stock and Vineyard Soil using Droplet Digital PCR

Plant Disease ◽  
2018 ◽  
Vol 102 (11) ◽  
pp. 2136-2141 ◽  
Author(s):  
Tanja M. Voegel ◽  
Louise M. Nelson
Keyword(s):  
Management Strategies ◽  
Digital Pcr ◽  
Agrobacterium Vitis ◽  
Nursery Stock ◽  
Vineyard Soil ◽  
Polymerase Chain ◽  
Soil Information ◽  
Digital Polymerase Chain Reaction ◽  
Current Detection ◽  
Infected Grapevine

Current detection methodologies for Agrobacterium vitis, causing crown gall of grapevines, are time intensive and lack the ability to quantify pathogen abundance in nursery stock and soil. Information on pathogen abundance is a key component to develop management strategies. The aim of this study was to develop a rapid and sensitive quantification assay for grapevine nursery stock and vineyard soil via droplet digital polymerase chain reaction targeting the virA gene. DNA isolated from roots of dormant grapevines originating from nurseries in Germany, California, and Ontario were tested for virA abundance. Bacterial numbers varied with grapevine origin; plants from California had the highest numbers. In addition, rhizosphere soil from two vineyards in the Okanagan valley in British Columbia was tested over a growing season. Sampling time during the season did not affect virA gene abundance. The older vineyard had higher soil A. vitis populations than the younger vineyard. The assay developed here has potential for use in national clean plant programs to prevent import of infected grapevine nursery stock and to test vineyard soil for abundance of the pathogen before planting.

scholarly journals Droplet Digital PCR (ddPCR) Analysis for the Detection and Quantification of Cow DNA in Buffalo Mozzarella Cheese

Animals ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 1270
Author(s):  
Anna Cutarelli ◽  
Andrea Fulgione ◽  
Pasquale Fraulo ◽  
Francesco Paolo Serpe ◽  
Pasquale Gallo ◽  
...  
Keyword(s):  
Digital Pcr ◽  
Buffalo Milk ◽  
Cow Milk ◽  
Mozzarella Cheese ◽  
Chain Reaction ◽  
New Methods ◽  
Protected Designation Of Origin ◽  
Commission Regulation ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

Buffalo mozzarella cheese is one of the most appreciated traditional Italian products and it is certified as a Protected Designation of Origin (PDO) product under the European Commission Regulation No. 1151/2012. It is obtained exclusively from buffalo milk. If made from cow milk, or a mixture of buffalo and cow milk, buffalo mozzarella cheese does not qualify as a PDO product. In order to maximize their profits, some producers market buffalo mozzarella that also contains cow milk as a PDO product, thus defrauding consumers. New methods for revealing this fraud are therefore needed. One such method is the droplet digital Polymerase Chain Reaction (ddPCR). Thanks to its high precision and sensitivity, the ddPCR could prove an efficacious means for detecting the presence of cow milk in buffalo mozzarella cheese that is marketed as a PDO product. ddPCR has proved able to detect the DNA of cow and/or buffalo milk in 33 buffalo mozzarella cheeses labelled as PDO products, and experimental evidence could support its application in routine analyses.

scholarly journals Clarity Plus™ digital PCR: A novel platform for absolute quantification of SARS-CoV-2

2021 ◽  
Author(s):  
Shawn Yi Han Tan ◽  
Milton Sheng Yi Kwek ◽  
Huiyu Low ◽  
Yan Ling Joy Pang
Keyword(s):  
Dynamic Range ◽  
Digital Pcr ◽  
Absolute Quantification ◽  
Viral Loads ◽  
Nucleic Acid Analysis ◽  
Polymerase Chain ◽  
The Absolute ◽  
Assay Precision ◽  
Digital Polymerase Chain Reaction ◽  
Higher Sensitivity

In recent years, the usage of digital polymerase chain reaction (dPCR) for various clinical applications has increased exponentially. Considering the growing demand for improved dPCR technology, the Clarity Plus™ dPCR system which features enhanced multiplexing capability and a wider dynamic range for nucleic acid analysis was recently launched. In this study, a dPCR assay optimized for use on Clarity Plus™ was evaluated for the absolute quantification of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent responsible for the global coronavirus disease 2019 (COVID-19) outbreak. The assay demonstrated good inter- and intra- assay precision, accuracy, as well as excellent linearity across a range of over 6 orders of magnitude for target gene quantification. In addition, comparison of the assay on both dPCR and qPCR platforms revealed that dPCR exhibited a slightly higher sensitivity compared to its qPCR counterpart when quantifying SARS-CoV-2 at a lower concentration. Overall, the results showed that the dPCR assay is a reliable and effective approach for the absolute quantification of SARS-CoV-2 and can potentially be adopted as a molecular tool in applications such as detecting low viral loads in patients as well as in wastewater surveillance of COVID-19.

scholarly journals ddpcr: an R package and web application for analysis of droplet digital PCR data

F1000Research ◽  
2016 ◽  
Vol 5 ◽  
pp. 1411 ◽  
Author(s):  
Dean Attali ◽  
Roza Bidshahri ◽  
Charles Haynes ◽  
Jennifer Bryan
Keyword(s):  
Web Application ◽  
R Package ◽  
Digital Pcr ◽  
Clinical Diagnostics ◽  
Great Promise ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction ◽  
Digital Nature ◽  
Accurate Quantification ◽  
Higher Sensitivity

Droplet digital polymerase chain reaction (ddPCR) is a novel platform for exact quantification of DNA which holds great promise in clinical diagnostics. It is increasingly popular due to its digital nature, which provides more accurate quantification and higher sensitivity than traditional real-time PCR. However, clinical adoption has been slowed in part by the lack of software tools available for analyzing ddPCR data. Here, we present ddpcr – a new R package for ddPCR visualization and analysis. In addition, ddpcr includes a web application (powered by the Shiny R package) that allows users to analyze ddPCR data using an interactive graphical interface.

scholarly journals Detection of fish allergen by droplet digital PCR

2019 ◽  
Vol 7 (4) ◽  
Author(s):  
Cinzia Daga ◽  
Simona Cau ◽  
Maria Giovanna Tilocca ◽  
Barbara Soro ◽  
Aldo Marongiu ◽  
...  
Keyword(s):  
18S Rrna ◽  
Annealing Temperature ◽  
Pcr Primers ◽  
Digital Pcr ◽  
18S Rrna Gene ◽  
Rrna Gene ◽  
Chain Reaction ◽  
European Regulation ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

Fish is one of fourteen allergens that must be highlighted on the label within the ingredients list. It should be noted that the European regulation, is very restrictive to allergens with zero tolerance. Therefore it is important to establish sensitive and specific methods for detecting fish allergen. Applicability to detect and quantify fish allergen by droplet digital polymerase chain reaction (ddPCR) has been evaluated in this work. Genomic DNA of three fish species belonging to the most common fish families were analyzed. PCR primers were designed to amplify a 166 bp region of the 18S rRNA gene. Comparative studies were performed to establish the optimal primer and probe concentrations.  Annealing temperature was determined by using thermal gradient. The results have shown good applicability of the optimized 18S rRNA gene-method to detect and quantify small amounts of the target in all samples analyzed. However, validation studies are needed in order to apply ddPCR technology for routine allergens analysis.  

scholarly journals Digital PCR: A Reliable Tool for Analyzing and Monitoring Hematologic Malignancies

2020 ◽  
Vol 21 (9) ◽  
pp. 3141 ◽  
Author(s):  
Nicoletta Coccaro ◽  
Giuseppina Tota ◽  
Luisa Anelli ◽  
Antonella Zagaria ◽  
Giorgina Specchia ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Residual Disease ◽  
Hematologic Malignancies ◽  
Digital Pcr ◽  
Future Perspectives ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction ◽  
Generation Sequencing ◽  
Minimal Residual

The digital polymerase chain reaction (dPCR) is considered to be the third-generation polymerase chain reaction (PCR), as it yields direct, absolute and precise measures of target sequences. dPCR has proven particularly useful for the accurate detection and quantification of low-abundance nucleic acids, highlighting its advantages in cancer diagnosis and in predicting recurrence and monitoring minimal residual disease, mostly coupled with next generation sequencing. In the last few years, a series of studies have employed dPCR for the analysis of hematologic malignancies. In this review, we will summarize these findings, attempting to focus on the potential future perspectives of the application of this promising technology.

Heterogeneous modification of through-hole microwell chips for ultralow cross-contamination digital polymerase chain reaction

The Analyst ◽  
2020 ◽  
Vol 145 (8) ◽  
pp. 3116-3124
Author(s):  
Jinze Li ◽  
Yajun Qiu ◽  
Zhiqi Zhang ◽  
Chuanyu Li ◽  
Shuli Li ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Digital Pcr ◽  
Cross Contamination ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction ◽  
Through Hole

Heterogeneous modification of through-hole microwell chips to avoid cross-contamination during digital PCR.

scholarly journals Application of droplet digital PCR to detect the pathogens of infectious diseases

2018 ◽  
Vol 38 (6) ◽  
Author(s):  
Haiyi Li ◽  
Ruolan Bai ◽  
Zhenyu Zhao ◽  
Lvyan Tao ◽  
Mingbiao Ma ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Infectious Diseases ◽  
Quantitative Polymerase Chain Reaction ◽  
Digital Pcr ◽  
Conventional Polymerase Chain Reaction ◽  
Nucleic Acid Detection ◽  
Chain Reaction ◽  
Molecular Biology Technique ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

Polymerase chain reaction (PCR) is a molecular biology technique used to multiply certain deoxyribonucleic acid (DNA) fragments. It is a common and indispensable technique that has been applied in many areas, especially in clinical laboratories. The third generation of polymerase chain reaction, droplet digital polymerase chain reaction (ddPCR), is a biotechnological refinement of conventional polymerase chain reaction methods that can be used to directly quantify and clonally amplify DNA. Droplet digital polymerase chain reaction is now widely used in low-abundance nucleic acid detection and is useful in diagnosis of infectious diseases. Here, we summarized the potential advantages of droplet digital polymerase chain reaction in clinical diagnosis of infectious diseases, including viral diseases, bacterial diseases and parasite infections, concluded that ddPCR provides a more sensitive, accurate, and reproducible detection of low-abundance pathogens and may be a better choice than quantitative polymerase chain reaction for clinical applications in the future.

A rapid nucleic acid concentration measurement system with large-field-of-view for droplet digital PCR microfluidic chip

Lab on a Chip ◽  
2021 ◽  
Author(s):  
Jinrong Shen ◽  
Jihong Zheng ◽  
Zhenqing Li ◽  
Yourong Liu ◽  
Fengxiang Jing ◽  
...  
Keyword(s):  
Digital Pcr ◽  
Absolute Quantification ◽  
Droplet Digital Pcr ◽  
Field Of View ◽  
Large Field ◽  
Chain Reaction ◽  
Effective Technique ◽  
Nucleic Acid Concentration ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

Droplet digital polymerase chain reaction(ddPCR) is an effective technique for the absolute quantification of target mucleic acid unparalleled sensitivity. However, current commerical ddPCR device for the detection of the gene...

scholarly journals A New Strategy for the Detection of Chicken Infectious Anemia Virus Contamination in Attenuated Live Vaccine by Droplet Digital PCR

2019 ◽  
Vol 2019 ◽  
pp. 1-9
Author(s):  
Qiuchen Li ◽  
Yubiao Zhang ◽  
Fanfeng Meng ◽  
Hui Jiang ◽  
Guanlong Xu ◽  
...  
Keyword(s):  
Digital Pcr ◽  
Economic Losses ◽  
Lymphoid Tissues ◽  
Detection Technology ◽  
Chicken Infectious Anemia Virus ◽  
New Strategy ◽  
Chicken Infectious Anemia ◽  
Polymerase Chain ◽  
Anemia Virus ◽  
Digital Polymerase Chain Reaction

Chicken infectious anemia virus (CIAV) causes the atrophy of bone marrow hematopoietic and lymphoid tissues in chicks, leading to huge economic losses all over the world. The using of attenuated vaccine contaminated with CIAV increased the mortality and the pathogenicity of other diseases in many farms. However, it is difficult to detect the CIAV contamination by general detection technology due to the extremely low dose of CIAV in vaccines. In this study, we established a new method called droplet digital Polymerase Chain Reaction (ddPCR) to detect CIAV contamination of vaccines more sensitively and accurately. The lowest detection limitation of this method is 2.4 copies of CIAV plasmid or CIAV contamination at 0.1 EID50/1000 feathers in vaccines without any positive signals of other viruses. Besides, the sensitivity of ddPCR is 100 times greater than that of conventional PCR and 10 times greater than that of real-time PCR. The ddPCR technique is more sensitive and more intuitive. Therefore, it could be valuable for the detection of CIAV contamination in vaccines.

A localized temporary negative pressure assisted microfluidic device for detecting keratin 19 in A549 lung carcinoma cells with digital PCR

2015 ◽  
Vol 7 (5) ◽  
pp. 2006-2011 ◽  
Author(s):  
Qingchang Tian ◽  
Qi Song ◽  
Yanan Xu ◽  
Qiangyuan Zhu ◽  
Bingwen Yu ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Digital Pcr ◽  
Carcinoma Cells ◽  
Biological Research ◽  
Chain Reaction ◽  
Lung Carcinoma Cells ◽  
Keratin 19 ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction ◽  
A549 Lung

Digital polymerase chain reaction (dPCR) has played a major role in biological research, especially by providing an accurate counting of single nucleic acid molecules.

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