Development of a Multiplex PCR Assay to Monitor Living Modified Cottons in South Korea

Cotton has been cultivated worldwide and is a useful crop for humans. However, all living modified organisms (LMOs), including living modified (LM) cotton, are not cultivated in South Korea and are imported from overseas. LM cotton imports are on the rise and most of the imported cotton is used as livestock feed. In particular, it is commonly used to feed Holstein breeds that produce milk, because cotton improves the quality of milk. However, as the cotton imports increase, the possibility of unintentional outflows in the distribution process also increases. Consequently, there is an increasing concern about unintentional release of LM cotton into the natural environment. Therefore, environmental monitoring and post-management of LMOs are very important steps. Recently, a total of 30 LM crop events were approved for LM cotton import in South Korea. A single detection method has been used to monitor individual events. However, a single method of detection for collected samples requires a large number of PCRs, with obvious disadvantages. Therefore, a simultaneous detection method was developed for 8 representative events (GHB119, GHB614, MON88913, MON15985, LLCOTTON25, MON1445, 281-3006, and MON531) in an effort to monitor 26 of them and facilitate the identification of LM cotton. The results suggest that our new multiplex PCR method may be useful for monitoring and post-management of LM cotton.

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Monitoring Living Modified Canola Using an Efficient Multiplex PCR Assay in Natural Environments in South Korea

Applied Sciences ◽

10.3390/app10217721 ◽

2020 ◽

Vol 10 (21) ◽

pp. 7721

Author(s):

Il Ryong Kim ◽

Hye Song Lim ◽

Wonkyun Choi ◽

Da In Kang ◽

Sang Yeol Lee ◽

...

Keyword(s):

South Korea ◽

Multiplex Pcr ◽

Natural Environments ◽

Pcr Assay ◽

Brassica Napus L ◽

Livestock Feed ◽

Multiplex Pcr Assay ◽

Management Project ◽

Pcr Method ◽

Living Modified Organisms

Canola (Brassica napus L.) is cultivated worldwide and utilized as a vegetable oil, biodiesel, and livestock feed. It is also a major living modified (LM) crop alongside corn, soybean, and cotton. Many canola events have been authorized for food, feed, and processing use in South Korea. Concerns about the unintentional release of LM canola into the natural environment have increased environmental monitoring and post-management of living modified organisms (LMOs) is on the rise. The Ministry of Environment (MOE) and the National Institute of Ecology (NIE) conducted an environmental LMO monitoring and post-management project for LM canola from 2014 to 2017. The number of suspicious LM samples gradually increased each year. In this study, a multiplex PCR method was established to detect seven single LM canola events (Topas 19/2, Rf3, Dp-73496-4, Ms8, GT73, Mon88032, and T45) to cover 14 approved LM canola events. This method was utilized to detect 22 LMs out of 260 suspicious canola samples. Thus, this new method is more efficient in terms of time and cost than conventional PCR methods for the identification and monitoring of LMOs.

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Multiplex PCR Method for the Simultaneous Detection of Histamine-, Tyramine-, and Putrescine-Producing Lactic Acid Bacteria in Foods

Journal of Food Protection ◽

10.4315/0362-028x-68.4.874 ◽

2005 ◽

Vol 68 (4) ◽

pp. 874-878 ◽

Cited By ~ 58

Author(s):

ÁNGELA MARCOBAL ◽

BLANCA de las RIVAS ◽

M. VICTORIA MORENO-ARRIBAS ◽

ROSARIO MUÑOZ

Keyword(s):

Lactic Acid ◽

Lactic Acid Bacteria ◽

Multiplex Pcr ◽

Simultaneous Detection ◽

Fermented Foods ◽

Multiplex Pcr Assay ◽

Pcr Method ◽

Dna Fragment ◽

Optimized Conditions ◽

Selection Of

In a screening of primers, we have selected three pairs of primers for a multiplex PCR assay for the simultaneous detection of lactic acid bacteria (LAB) strains, which potentially produce histamine, tyramine, and putrescine on fermented foods. These primers were based on sequences from histidine, tyrosine, and ornithine decarboxylases from LAB. Under the optimized conditions, the assay yielded a 367-bp DNA fragment from histidine decarboxylases, a 924-bp fragment from tyrosine decarboxylases, and a 1,446-bp fragment from ornithine decarboxylases. When the DNAs of several target organisms were included in the same reaction, two or three corresponding amplicons of different sizes were observed. This assay was useful for the detection of amine-producing bacteria in control collection strains and in a LAB collection. No amplification was observed with DNA from nonproducing LAB strains. This article is the first describing a multiplex PCR approach for the simultaneous detection of potentially amine-producing LAB in foods. It can be easily incorporated into the routine screening for the accurate selection of starter LAB and in food control laboratories.

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A PCR method targeting internal transcribed spacers: the simultaneous detection of Babesia bigemina and Babesia bovis in cattle

Acta Parasitologica ◽

10.2478/s11686-014-0222-6 ◽

2014 ◽

Vol 59 (1) ◽

Cited By ~ 8

Author(s):

Junlong Liu ◽

Guiquan Guan ◽

Aihong Liu ◽

Youquan Li ◽

Hong Yin ◽

...

Keyword(s):

Light Microscopy ◽

Multiplex Pcr ◽

Simultaneous Detection ◽

Internal Transcribed Spacers ◽

Babesia Bovis ◽

Babesia Bigemina ◽

Pcr Assay ◽

Blood Samples ◽

Multiplex Pcr Assay ◽

Pcr Method

AbstractIn this study, two pairs of oligonucleotide primers were designed according to the nucleotide sequence of the internal transcribed spacers (ITSs) of Babesia bigemina and B. bovis isolates from China. The primers were used in a multiplex PCR to detect parasite DNA in blood samples from cattle. There was no cross reactions with B. ovata, B. major, B. sp. Kashi, Theileria annulata, T. sergenti, T. sinensis or normal bovine DNA. The sensitivity of multiplex PCR assay was 1 pg and 10 pg DNA for B. bigemina and B. bovis, respectively. A total of 260 field blood samples collected from cattle in five provinces of China were analyzed by multiplex PCR and light microscopy. PCR testing revealed that 7.3% (19/260) and 5.8% (15/260) of cattle were positive for B. bigemina and B. bovis and 1.2% (3/260) of cattle were co-infected with B. bigemina and B. bovis. Using light microscopy, 2.3% (6/260) and 1.5% (4/260) of cattle were infected by B. bigemina and B. bovis, respectively, and no co-infection was found. The results showed that the multiplex PCR developed in the present study could be an alternative diagnostic tool for the detection of B. bovis and B. bigemina infection in cattle.

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A novel common primer multiplex PCR (CP-M-PCR) method for the simultaneous detection of meat species

Food Control ◽

10.1016/j.foodcont.2008.05.021 ◽

2009 ◽

Vol 20 (4) ◽

pp. 366-370 ◽

Cited By ~ 38

Author(s):

Weibin Bai ◽

Wentao Xu ◽

Kunlun Huang ◽

Yanfang Yuan ◽

Sishuo Cao ◽

...

Keyword(s):

Multiplex Pcr ◽

Simultaneous Detection ◽

Meat Species ◽

Pcr Method

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Development and evaluation of a multiplex PCR assay for simultaneous detection ofFlavobacterium psychrophilum,Yersinia ruckeriandAeromonas salmonicidasubsp.salmonicidain culture fisheries

Journal of Veterinary Science ◽

10.4142/jvs.2010.11.3.235 ◽

2010 ◽

Vol 11 (3) ◽

pp. 235 ◽

Cited By ~ 6

Author(s):

Ertan Emek Onuk ◽

Alper Ciftci ◽

Arzu Findik ◽

Yuksel Durmaz

Keyword(s):

Multiplex Pcr ◽

Simultaneous Detection ◽

Pcr Assay ◽

Multiplex Pcr Assay

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A new one-step multiplex PCR assay for simultaneous detection and identification of avian haemosporidian parasites

Parasitology Research ◽

10.1007/s00436-018-6153-7 ◽

2018 ◽

Vol 118 (1) ◽

pp. 191-201 ◽

Cited By ~ 17

Author(s):

Arif Ciloglu ◽

Vincenzo A. Ellis ◽

Rasa Bernotienė ◽

Gediminas Valkiūnas ◽

Staffan Bensch

Keyword(s):

Multiplex Pcr ◽

Simultaneous Detection ◽

Pcr Assay ◽

Haemosporidian Parasites ◽

Multiplex Pcr Assay ◽

Detection And Identification ◽

One Step

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Multiplex PCR Detection of Enterotoxin Genes in Aeromonas spp. from Suspect Food Samples in Northern Taiwan

Journal of Food Protection ◽

10.4315/0362-028x-71.10.2094 ◽

2008 ◽

Vol 71 (10) ◽

pp. 2094-2099 ◽

Cited By ~ 14

Author(s):

YU-CHANG CHANG ◽

JAN-YI WANG ◽

AMMAIYAPPAN SELVAM ◽

SHU-CHEN KAO ◽

SHANG-SHYNG YANG ◽

...

Keyword(s):

Multiplex Pcr ◽

Diarrheal Disease ◽

Simultaneous Detection ◽

Food Poisoning ◽

Food Samples ◽

Pcr Assay ◽

Multiplex Pcr Assay ◽

Enterotoxin Genes ◽

Causative Agents ◽

Northern Taiwan

Aeromonads possess an array of virulence factors and are causative agents of a number of human infections. Among them, genes of one cytotoxic (Act) and two cytotonic (Alt, Ast) enterotoxins are implicated in a human diarrheal disease. A rapid, specific, simultaneous detection of these enterotoxin genes in suspected food poisoning samples is not yet reported. Hence, a multiplex PCR assay was designed to amplify the cytotoxic (act), heat-labile cytotonic (alt), and heat-stable cytotonic (ast) enterotoxin genes of aeromonads. The PCR assay was tested with 133 Aeromonas spp. isolated from suspect food poisoning samples and retail samples of poultry and fish from wet markets in and around Taipei, Northern Taiwan. The Aeromonas spp. isolates were divided into six genotypes based on absence or presence of one or more enterotoxin genes. Of these 133 isolates, Aeromonas caviae (52.5%) and Aeromonas hydrophila (43.4%) were the most frequently isolated species from food poisoning samples and retail samples, respectively. Among the species, A. hydrophila had a significantly higher proportion for harboring three enterotoxin genes than had the others, whereas Aeromonas encheleia, considered a nonpathogen, was found harboring three enterotoxin genes. The multiplex PCR assays are rapid and specific, and provide a useful tool for the detection and genotyping of enterotoxin genes of aeromonads.

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