Real-time loop-mediated isothermal amplification for the CaMV-35S promoter as a screening method for genetically modified organisms

Fukuta, Shiro; Mizukami, Yuko; Ishida, Akira; Ueda, Jun-ichi; Hasegawa, Mayumi; Hayashi, Izumi; Hashimoto, Minako; Kanbe, Michio

doi:10.1007/s00217-003-0862-5

Cited by 64 publications

(41 citation statements)

References 17 publications

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“…Therefore, the size of target DNA should be set to less than 300 bp, including FIP (FIP-cry and FIP-chi) and BIP (BIP-cry and BIP-chi). Most of the LAMP methods used for GMO detection generally focus on the sequence of frequently used genetic elements, such as promoters, terminators (Fukuta et al 2004;Lee et al 2009a), marker genes (Guan et al 2010), etc. However, LAMP primers can also be easily adapted to other target genes.…”

Section: Discussionmentioning

confidence: 99%

“…Because of the specific features of LAMP primers, target selectivity is expected to be higher than that in PCR (Notami et al 2000). High specificity is assured since target recognition is mediated by four distinct primers targeting six different sequences (Notami et al 2000;Nagamine et al 2002;Fukuta et al 2004). In some studies, LAMP was even shown to be more selective (Seki et al 2005) and less sensitive to background DNA (Notami et al 2000) than PCR.…”

Section: Discussionmentioning

confidence: 99%

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Rapid identification of transgenic cotton (Gossypium hirsutum L.) plants by loop-mediated isothermal amplification

Rostamkhani¹,

Haghnazari²,

Tohidfar³

et al. 2011

CZECH J GENET PLANT

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Abstract:In an attempt to speed up the process of screening of transgenic cotton (G. hirsutum L.) plants, a visual and rapid loop-mediated isothermal amplification (LAMP) assay was adopted. Genomic DNA was extracted from fresh leaf tissues of T 2 transgenic cotton containing chitinase (chi) and cry1A(b) genes. Detection of genes of interest was performed by polymerase chain reaction (PCR), LAMP and real-time PCR methods. In LAMP assay the amplification was performed after 30 min at 65°C when loop primers were involved in the reaction. The involvement of loop primers decreased the time needed for amplification. By testing serial tenfold dilutions (10 -1 to 10 -8 ) of the genes of interest, the detection sensitivity of LAMP was found to be 100-fold higher than that of PCR. The rapid DNA extraction method and LAMP assay can be performed within 30 min and the derived LAMP products can be directly observed as visually detectable based on turbidity in the reaction tube. The accuracy of LAMP method in the screening of transgenes was confirmed by PCR and real-time PCR. The developed method was efficient, rapid and sensitive in the screening of cotton transgenic plants. This method can be applied to any other crops.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Rapid identification of transgenic cotton (Gossypium hirsutum L.) plants by loop-mediated isothermal amplification

Rostamkhani¹,

Haghnazari²,

Tohidfar³

et al. 2011

CZECH J GENET PLANT

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“…A GMO screening method using LAMP targeting GMelement Cauliflower Mosaic Virus 35S (P-35S) was first reported by Fukuta et al (2004) [22]. So far, LAMP methods for T-nos, P-35S, P-nos [23] and the pat marker gene [24] have been developed for GMO element screening in the laboratory.…”

Section: The Screening Of Gmomentioning

confidence: 99%

Loop-Mediated Isothermal Amplification (LAMP) Assay for GMO on: Recent Progresses and Future Perspectives

Li¹,

Ji²,

Zhao³

et al. 2015

OALib

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As more and more genetically modified (GM) crops are approved for commercialization and planting, safety issues of GM crops have become hot topics worldwide. For both regulatory and academic purposes, development of rapid, economic and effective on-site detection methods for GM components is indispensible. Up to now, the most effective and sensitive techniques used for GMO detection are based on polymerase chain reaction (PCR). PCR method needs expensive, heavy instruments and gel electrophoresis. Therefore, it is commonly used in laboratory test, and unsuitable for on-site detection. Loop-mediated isothermal amplification (LAMP), an isothermal nucleic acid amplification technique, has been extensively used in many areas such as food safety and clinic diagnosis. Advantageous characteristics of LAMP, such as high specificity and sensitivity, simple operation, low cost, eye visualization, particularly free of special equipment, render it with high potential to be used for GMO on-site detection. In this review, we summarized current status of the application of LAMP in GMO detection, and discussed possible improvements needed for its adaptability regarding to on-site GMO detection. Hopefully, the information present here would facilitate the practical risk assessment of GMO.

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“…Due to the higher specificity, sensitivity and amplification efficiency, LAMP had been widely used for the diagnosis of viruses (Bista et al, 2007;Yoneyama et al, 2007) and pathogenic microorganisms (Iwamoto et al, 2003;Kuboki et al, 2003;HaraKudo et al, 2008), but it can hardly be seen on the application for GM crops up to now (Fukuta et al, 2004;Lee et al, 2009;Guan et al, 2010;Chen et al, 2011).…”

Section: Introductionmentioning

confidence: 99%

Development of the One-step Visual Loop-Mediated Isothermal Amplification Assay for Genetically Modified Rice Event TT51-1

Chen

Wang

Zhu

et al. 2014

FSTR

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Loop-mediated isothermal amplification (LAMP), a novel DNA amplification technique, is widely used inmolecular diagnosis although it is very easy to be contaminated by the huge amount of its own amplicons. An improved visual LAMP method for the event specific identification of GM rice TT51-1 and a rice endogenous reference gene (sucrose phosphate synthase, SPS) was established. Pre-loaded calcein and manganous ion into the LAMP reaction offered not only an easy way to read results by colour change from orange to green but also a closedtube system to reduce contaminations caused by amplification products. The LAMP assays could be finished within 60 min, and the limits of detection (LODs) were estimated as low as 10 and 20 copies of rice haploid genomic DNA, respectively, about 10-fold more sensitive than that of conventional PCR. The specificities of the assays were also well evaluated with optimized system and conditions. The developed one-step visual LAMP assays in this study provided a promising tool for the convenient and effective detection of GM crops.

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Real-time loop-mediated isothermal amplification for the CaMV-35S promoter as a screening method for genetically modified organisms

Cited by 64 publications

References 17 publications

Rapid identification of transgenic cotton (Gossypium hirsutum L.) plants by loop-mediated isothermal amplification

Rapid identification of transgenic cotton (Gossypium hirsutum L.) plants by loop-mediated isothermal amplification

Loop-Mediated Isothermal Amplification (LAMP) Assay for GMO on: Recent Progresses and Future Perspectives

Development of the One-step Visual Loop-Mediated Isothermal Amplification Assay for Genetically Modified Rice Event TT51-1

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