Browse
Submit Data
Databases
API
Help

Dataset Information

16 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Development of a rapid detection method for genetically modified rice using the ultra-fast PCR system.

ABSTRACT: Genetically modified (GM) rice varieties containing traits such as tolerance to abiotic stress and resistance against pests and diseases continue to be developed. However, contamination incidents from unauthorized GM rice varieties have been encountered. To date, no GM rice crop has been authorized for consumption and/or commercialization in Korea. Therefore, to enhance safety management of unauthorized genetically modified organisms (GMOs), accurate and reliable detection methods are needed to identify GMOs in crops or products. In this study, we developed rapid detection methods for GM rice events (Bt63, KMD1, Kefeng6, Kefeng8, and LLRice62) using ultra-fast PCR system. Ultra-fast PCR is a state-of-the-art technology and decreases PCR run-times dramatically. However, the ultra-fast PCR is not widely used in GMO analysis. Thus, we designed a detection method for five events of GM rice and confirmed them by performing specificity, sensitivity, and applicability assays. All results demonstrate that the ultra-fast PCR system is a specific, sensitive, and reliable method to identify and monitor GM rice events. Additionally, it can be utilized as a rapid and simple method for GMO analysis in crops or processed products. This study can be used as a reference for future research on new analysis methods of unauthorized GMOs.

SUBMITTER: Shin MK

PROVIDER: S-EPMC8783574 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Detection method for genetically modified potato using an ultra-fast PCR system.

Project description:Genetically modified (GM) potatoes having resistance to insects and viral diseases, low reducing sugar contents, and black spots for high quality continue to be developed. However, no GM potato has been approved as food or feed in the Republic of Korea as the country adheres to a zero-tolerance policy to unauthorized genetically modified organisms (GMOs). When the self-sufficiency rate is low, a detection method to assess GMOs in crops or other products is necessary. Therefore, a rapid method for two GM potato events (SPS-Y9 and EH92-527-1) using an ultra-fast PCR (UF-PCR) system has been developed, and its specificity, sensitivity, and applicability were demonstrated. UF-PCR can decrease the runtime of PCR by more than half of that needed in conventional methods. However, UF-PCR is not a common method for GMO analysis. This rapid detection method may be useful for GMO analyses in field conditions.

| S-EPMC9891748 | biostudies-literature

Rapid Monitoring of Viable Genetically Modified Escherichia coli Using a Cell-Direct Quantitative PCR Method Combined with Propidium Monoazide Treatment

Project description:The commercialization of industrial genetically modified microorganisms (GMMs) has highlighted their impact on public health and the environment. Rapid and effective monitoring methods detecting live GMMs are essential to enhance current safety management protocols. This study aims to develop a novel cell-direct quantitative polymerase chain reaction (qPCR) method targeting two antibiotic-resistant genes, KmR and nptII, conferring resistance against kanamycin and neomycin, along with propidium monoazide, to precisely detect viable Escherichia coli. The E. coli single-copy taxon-specific gene of D-1-deoxyxylulose 5-phosphate synthase (dxs) was used as the internal control. The qPCR assays demonstrated good performance, with dual-plex primer/probe combinations exhibiting specificity, absence of matrix effects, linear dynamic ranges with acceptable amplification efficiencies, and repeatability for DNA, cells, and PMA-treated cells targeting KmR/dxs and nptII/dxs. Following the PMA-qPCR assays, the viable cell counts for KmR-resistant and nptII-resistant E. coli strains exhibited a bias% of 24.09% and 0.49%, respectively, which were within the acceptable limit of ±25%, as specified by the European Network of GMO Laboratories. This method successfully established detection limits of 69 and 67 viable genetically modified E. coli cells targeting KmR and nptII, respectively. This provides a feasible monitoring approach as an alternative to DNA processing techniques to detect viable GMMs.

| S-EPMC10220751 | biostudies-literature

A highly sensitive and specific method for the screening detection of genetically modified organisms based on digital PCR without pretreatment.

Project description:Digital PCR has developed rapidly since it was first reported in the 1990 s. It was recently reported that an improved method facilitated the detection of genetically modified organisms (GMOs). However, to use this improved method, the samples must be pretreated, which could introduce inaccuracy into the results. In our study, we explored a pretreatment-free digital PCR detection method for the screening for GMOs. We chose the CaMV35s promoter and the NOS terminator as the templates in our assay. To determine the specificity of our method, 9 events of GMOs were collected, including MON810, MON863, TC1507, MIR604, MIR162, GA21, T25, NK603 and Bt176. Moreover, the sensitivity, intra-laboratory and inter-laboratory reproducibility of our detection method were assessed. The results showed that the limit of detection of our method was 0.1%, which was lower than the labeling threshold level of the EU. The specificity and stability among the 9 events were consistent, respectively. The intra-laboratory and inter-laboratory reproducibility were both good. Finally, the perfect fitness for the detection of eight double-blind samples indicated the good practicability of our method. In conclusion, the method in our study would allow more sensitive, specific and stable screening detection of the GMO content of international trading products.

| S-EPMC4530665 | biostudies-other

RT-RPA-PfAgo System: A Rapid, Sensitive, and Specific Multiplex Detection Method for Rice-Infecting Viruses.

Project description:The advancement in CRISPR-Cas biosensors has transmuted the detection of plant viruses owing to their rapid and higher sensitivity. However, false positives and restricted multiplexing capabilities are still the challenges faced by this technology, demanding the exploration of novel methodologies. In this study, a novel detection system was developed by integrating reverse transcriptome (RT) techniques with recombinase polymerase isothermal amplification (RPA) and Pyrococcus furiosus Argonaute (PfAgo). The RT-RPA-PfAgo system enabled the simultaneous detection of rice ragged stunt virus (RRSV), rice grassy stunt virus (RGSV), and rice black streaked dwarf virus (RBSDV). Identifying targets via guide DNA without being hindered by protospacer adjacent motif sequences is the inherent merit of PfAgo, with the additional advantage of it being simple, cost-effective, and exceptionally sensitive, with detection limits between 3.13 and 5.13 copies/µL, in addition to it effectively differentiating between the three distinct viruses. The field evaluations were also in accordance with RT-PCR methods. The RT-RPA-PfAgo system proved to be a robust, versatile, highly specific, and sensitive method with great potential for practicality in future plant virus diagnostics.

| S-EPMC10605773 | biostudies-literature

A PCR detection method for rapid identification of Paenibacillus larvae.

Project description:American foulbrood is a disease of larval honeybees (Apis mellifera) caused by the bacterium Paenibacillus larvae. Over the years attempts have been made to develop a selective medium for the detection of P. larvae spores from honey samples. The most successful of these is a semiselective medium containing nalidixic acid and pipermedic acid. Although this medium allows the growth of P. larvae and prevents the growth of most other bacterial species, the false-positive colonies that grow on it prevent the rapid confirmation of the presence of P. larvae. Here we describe a PCR detection method which can be used on the colonies that grow on this semiselective medium and thereby allows the rapid confirmation of the presence of P. larvae. The PCR primers were designed on the basis of the 16S rRNA gene of P. larvae and selectively amplify a 973-bp amplicon. The PCR amplicon was confirmed as originating from P. larvae by sequencing in both directions. Detection was specific for P. larvae, and the primers did not hybridize with DNA from closely related bacterial species.

| S-EPMC91325 | biostudies-literature

A novel quadruplex real-time PCR method for simultaneous detection of Cry2Ae and two genetically modified cotton events (GHB119 and T304-40).

Project description:BACKGROUND: To date, over 150 genetically modified (GM) crops are widely cultivated. To comply with regulations developed for genetically modified organisms (GMOs), including labeling policies, many detection methods for GMO identification and quantification have been developed. RESULTS: To detect the entrance and exit of unauthorized GM crop events in China, we developed a novel quadruplex real-time PCR method for simultaneous detection and quantification of GM cotton events GHB119 and T304-40 in cotton-derived products (based on the 5'-flanking sequence) and the insect-resistance gene Cry2Ae. The limit of detection was 10 copies for GHB119 and Cry2Ae and 25 copies for T304-40. The limit of quantification was 25 copies for GHB119 and Cry2Ae and 50 copies for T304-40. Moreover, low bias and acceptable standard deviation and relative standard deviation values were obtained in quantification analysis of six blind samples containing different GHB119 and T304-40 ingredients. CONCLUSIONS: The developed quadruplex quantitative method could be used for quantitative detection of two GM cotton events (GHB119 and T304-40) and Cry2Ae gene ingredient in cotton derived products.

| S-EPMC4064275 | biostudies-literature

Endpoint visual detection of three genetically modified rice events by loop-mediated isothermal amplification.

Project description:Genetically modified (GM) rice KMD1, TT51-1, and KF6 are three of the most well known transgenic Bt rice lines in China. A rapid and sensitive molecular assay for risk assessment of GM rice is needed. Polymerase chain reaction (PCR), currently the most common method for detecting genetically modified organisms, requires temperature cycling and relatively complex procedures. Here we developed a visual and rapid loop-mediated isothermal amplification (LAMP) method to amplify three GM rice event-specific junction sequences. Target DNA was amplified and visualized by two indicators (SYBR green or hydroxy naphthol blue [HNB]) within 60 min at an isothermal temperature of 63 °C. Different kinds of plants were selected to ensure the specificity of detection and the results of the non-target samples were negative, indicating that the primer sets for the three GM rice varieties had good levels of specificity. The sensitivity of LAMP, with detection limits at low concentration levels (0.01%?0.005% GM), was 10- to 100-fold greater than that of conventional PCR. Additionally, the LAMP assay coupled with an indicator (SYBR green or HNB) facilitated analysis. These findings revealed that the rapid detection method was suitable as a simple field-based test to determine the status of GM crops.

| S-EPMC3509588 | biostudies-literature

Loop-mediated isothermal amplification (LAMP) method for detection of genetically modified maize T25.

Project description:The loop-mediated isothermal amplification (LAMP) assay indicates a potential and valuable means for genetically modified organism (GMO) detection especially for its rapidity, simplicity, and low cost. We developed and evaluated the specificity and sensitivity of the LAMP method for rapid detection of the genetically modified (GM) maize T25. A set of six specific primers was successfully designed to recognize six distinct sequences on the target gene, including a pair of inner primers, a pair of outer primers, and a pair of loop primers. The optimum reaction temperature and time were verified to be 65°C and 45 min, respectively. The detection limit of this LAMP assay was 5 g kg(-1) GMO component. Comparative experiments showed that the LAMP assay was a simple, rapid, accurate, and specific method for detecting the GM maize T25.

| S-EPMC3951539 | biostudies-literature

Ultra-fast PCR method for the distinguishing between <i>Miichthys miiuy</i> and <i>Sciaenops ocellatus</i>.

Project description:The mi-iuy croaker Miichthys miiuy has immense commercial value in the Republic of Korea. The red drum Sciaenops ocellatus is widely produced by aquaculture, although its price is approximately 25% that of M. miiuy. S. ocellatus has black spots on its tail, enabling it to be distinguished from M. miiuy based on appearance. However, identifying S. ocellatus after simple processing steps, such as skin removal and dicing, is difficult. Certain traders misrepresent and sell S. ocellatus as M. miiuy or cultured M. miiuy for illegal economical gain. Therefore, an accurate and rapid identification method is required to distinguish between M. miiuy and S. ocellatus in the field. Here, a method for rapid field identification was developed based on species-specific primers using a portable ultra-fast PCR instrument. The ultra-fast real-time PCR method can complete the entire analytical procedure, including DNA isolation, amplification, and detection, within 30 min, thus maintaining the accuracy of identifying M. miiuy and S. ocellatus products on site. Forty-nine commercial products were tested, and all samples were successfully identified. Thus, the developed method is rapid, efficient tool for ensuring consumer protection.Supplementary informationThe online version contains supplementary material available at 10.1007/s10068-021-00954-4.

| S-EPMC8423894 | biostudies-literature

Rapid detection of MDR-Mycobacterium tuberculosis using modified PCR-SSCP from clinical Specimens.

Project description:ObjectiveTo design a rapid test to detect the rifampin (RIF) and isoniazid (INH) resistant mutant based on polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) technique that analyzes the katG, rpoB genes.MethodsBiochemical test as well as IS6110 targeting PCR revealed 103 clinical samples were tuberculosis. To determine the susceptibility of isolates to anti TB drugs, the proportional method was used. Mutations presented within the amplified products of the katG, rpoB genes were evaluated by SSCP.ResultsUsing proportional method, 12 (11.6%) and 9 (8.7%) isolates were resistant respectively to INH and RIF and 9 (8.7%) isolates showed resistance to both drug (multi-drug resistant tuberculosis). Three (2.9%) multi-drug resistant tuberculosis and two INH resistant isolates were detected by the PCR-SSCP and sequencing. The sensitivity and specificity of PCR-SSCP for multi-drug resistant isolates were 33% and 100%, respectively.ConclusionsComplete agreement between SSCP and sequencing can indicate that resistance-associated mutations have occurred in other genes except our considered genes.

| S-EPMC4025311 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data