Development of a real-time microchip PCR system for portable plant disease diagnosis.

Rapid and accurate detection of plant pathogens in the field is crucial to prevent the proliferation of infected crops. Polymerase chain reaction (PCR) process is the most reliable and accepted method for plant pathogen diagnosis, however current conventional PCR machines are not portable and requir...

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Autores principales: Chiwan Koo, Martha Malapi-Wight, Hyun Soo Kim, Osman S Cifci, Vanessa L Vaughn-Diaz, Bo Ma, Sungman Kim, Haron Abdel-Raziq, Kevin Ong, Young-Ki Jo, Dennis C Gross, Won-Bo Shim, Arum Han
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Publicado: Public Library of Science (PLoS) 2013
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spelling oai:doaj.org-article:15328817e0944924a28db997c9fb144b2021-11-18T08:42:12ZDevelopment of a real-time microchip PCR system for portable plant disease diagnosis.1932-620310.1371/journal.pone.0082704https://doaj.org/article/15328817e0944924a28db997c9fb144b2013-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/24349341/?tool=EBIhttps://doaj.org/toc/1932-6203Rapid and accurate detection of plant pathogens in the field is crucial to prevent the proliferation of infected crops. Polymerase chain reaction (PCR) process is the most reliable and accepted method for plant pathogen diagnosis, however current conventional PCR machines are not portable and require additional post-processing steps to detect the amplified DNA (amplicon) of pathogens. Real-time PCR can directly quantify the amplicon during the DNA amplification without the need for post processing, thus more suitable for field operations, however still takes time and require large instruments that are costly and not portable. Microchip PCR systems have emerged in the past decade to miniaturize conventional PCR systems and to reduce operation time and cost. Real-time microchip PCR systems have also emerged, but unfortunately all reported portable real-time microchip PCR systems require various auxiliary instruments. Here we present a stand-alone real-time microchip PCR system composed of a PCR reaction chamber microchip with integrated thin-film heater, a compact fluorescence detector to detect amplified DNA, a microcontroller to control the entire thermocycling operation with data acquisition capability, and a battery. The entire system is 25 × 16 × 8 cm(3) in size and 843 g in weight. The disposable microchip requires only 8-µl sample volume and a single PCR run consumes 110 mAh of power. A DNA extraction protocol, notably without the use of liquid nitrogen, chemicals, and other large lab equipment, was developed for field operations. The developed real-time microchip PCR system and the DNA extraction protocol were used to successfully detect six different fungal and bacterial plant pathogens with 100% success rate to a detection limit of 5 ng/8 µl sample.Chiwan KooMartha Malapi-WightHyun Soo KimOsman S CifciVanessa L Vaughn-DiazBo MaSungman KimHaron Abdel-RaziqKevin OngYoung-Ki JoDennis C GrossWon-Bo ShimArum HanPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 8, Iss 12, p e82704 (2013)
institution DOAJ
collection DOAJ
language EN
topic Medicine
R
Science
Q
spellingShingle Medicine
R
Science
Q
Chiwan Koo
Martha Malapi-Wight
Hyun Soo Kim
Osman S Cifci
Vanessa L Vaughn-Diaz
Bo Ma
Sungman Kim
Haron Abdel-Raziq
Kevin Ong
Young-Ki Jo
Dennis C Gross
Won-Bo Shim
Arum Han
Development of a real-time microchip PCR system for portable plant disease diagnosis.
description Rapid and accurate detection of plant pathogens in the field is crucial to prevent the proliferation of infected crops. Polymerase chain reaction (PCR) process is the most reliable and accepted method for plant pathogen diagnosis, however current conventional PCR machines are not portable and require additional post-processing steps to detect the amplified DNA (amplicon) of pathogens. Real-time PCR can directly quantify the amplicon during the DNA amplification without the need for post processing, thus more suitable for field operations, however still takes time and require large instruments that are costly and not portable. Microchip PCR systems have emerged in the past decade to miniaturize conventional PCR systems and to reduce operation time and cost. Real-time microchip PCR systems have also emerged, but unfortunately all reported portable real-time microchip PCR systems require various auxiliary instruments. Here we present a stand-alone real-time microchip PCR system composed of a PCR reaction chamber microchip with integrated thin-film heater, a compact fluorescence detector to detect amplified DNA, a microcontroller to control the entire thermocycling operation with data acquisition capability, and a battery. The entire system is 25 × 16 × 8 cm(3) in size and 843 g in weight. The disposable microchip requires only 8-µl sample volume and a single PCR run consumes 110 mAh of power. A DNA extraction protocol, notably without the use of liquid nitrogen, chemicals, and other large lab equipment, was developed for field operations. The developed real-time microchip PCR system and the DNA extraction protocol were used to successfully detect six different fungal and bacterial plant pathogens with 100% success rate to a detection limit of 5 ng/8 µl sample.
format article
author Chiwan Koo
Martha Malapi-Wight
Hyun Soo Kim
Osman S Cifci
Vanessa L Vaughn-Diaz
Bo Ma
Sungman Kim
Haron Abdel-Raziq
Kevin Ong
Young-Ki Jo
Dennis C Gross
Won-Bo Shim
Arum Han
author_facet Chiwan Koo
Martha Malapi-Wight
Hyun Soo Kim
Osman S Cifci
Vanessa L Vaughn-Diaz
Bo Ma
Sungman Kim
Haron Abdel-Raziq
Kevin Ong
Young-Ki Jo
Dennis C Gross
Won-Bo Shim
Arum Han
author_sort Chiwan Koo
title Development of a real-time microchip PCR system for portable plant disease diagnosis.
title_short Development of a real-time microchip PCR system for portable plant disease diagnosis.
title_full Development of a real-time microchip PCR system for portable plant disease diagnosis.
title_fullStr Development of a real-time microchip PCR system for portable plant disease diagnosis.
title_full_unstemmed Development of a real-time microchip PCR system for portable plant disease diagnosis.
title_sort development of a real-time microchip pcr system for portable plant disease diagnosis.
publisher Public Library of Science (PLoS)
publishDate 2013
url https://doaj.org/article/15328817e0944924a28db997c9fb144b
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