Loop-mediated Isothermal Amplification for the Detection of CaMV35S Promoter in Genetically Modified Maize

doi:10.7668/hbnxb.2011.04.002

Abstract

Abstract: The objective is to develop a loop-mediated isothermal amplification method for detection of CaMV-35S promoter in Genetically Modified maize.Loop-mediated isothermal amplification(LAMP),a novel nucleic acid amplification method,was developed for the rapid detection of Genetically Modified maize.Cauliflower mosaic virus 35S(CaMV35S) promoter gene was amplified by a set of four primers that recognize six distinct sequences of the target.The optimized conditions for LAMP were studied using different Mg²⁺,dNTPs,Betaine and primers concentrations.Furthermore,the sensitivity of LAMP was tested comparing with PCR.The LAMP method can detect CaMV35S promoter from genetically modified maize and their test results were consistent with the results of conventional PCR method.LAMP assay results were found to be more sensitive than the conventional PCR.The LAMP assay is an extremely rapid,highly sensitive,specific method,and will be an effective tool for rapid detection of Genetically Modified maize.

Key words: Genetically modified maize, Loop-mediated isothermal amplification(LAMP), Detection method

CLC Number:

S513.03

CHEN Jin-song, HUANG Cong-lin, ZHANG Xiu-hai, WU Zhong-yi. Loop-mediated Isothermal Amplification for the Detection of CaMV35S Promoter in Genetically Modified Maize[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2011, 26(4): 8-14. doi: 10.7668/hbnxb.2011.04.002.

/ / Recommend / Download Citations

URL: http://www.hbnxb.net/EN/10.7668/hbnxb.2011.04.002

http://www.hbnxb.net/EN/Y2011/V26/I4/8

References

[1] Clive James. 2009年全球生物技术/转基因作物商业化发展态势[J].中国生物工程杂志,2010,(02):1-22.
[2] Matsuoka T,Kuribara H,Akiyama H. A multiplex PCR method of detecting combinant DNAs from five lines of genetically modified maize[J].Shokuhin Eiseigaku Zasshi,2001,(01):24-32.doi:10.3358/shokueishi.42.24.
[3] Hardegger M,Brodmann P,Herrmann A. Quantitative detection of the 35S promoter and the NOS terminator u sing quantitative competitive PCR[J].European Food Research and Technology,1999,(02):83-87.doi:10.1007/s002170050462.
[4] Hübner P,Studer E,Lüthy J. Quantitative competitive PCR for the detection of genetically modified organisms in food[J].Food Control,1999,(06):353-358.
[5] Studer E,Rhyner C,Lüthy J. Quantitative competitive PCR for the detection of genetically modified soybean and maize[J].ZEITSCHRIFT für LEBENSMITTEL-UNTERSUCHUNG UND-FORSCHUNG A,1998,(03):207-213.
[6] AhmedF E. Application of molecular biology to biomedicine and toxicology[J].Journal of Environmental Science and Health Part A:Environmental Science and Engineering and Toxic and Hazardous Substance Control,1995.1-51.
[7] Hernández M,Esteve T,Prat S. Development of real-time PCR systems based on SYBR(R) Green I,AmplifluorTM and TaqMan(R) technologies for specific quantitative detection of the transgenic maize event GA21[J].Journal of Cereal Science,2004,(01):99-107.doi:10.1016/S0733-5210(03)00071-7.
[8] Chaouachi M,El Malki R,Berard A. Development of a real-time PCR method for the differential detection and quantification of four solanaceae in GMO analysis:potato (Solanum tuberosum),tomato (Solanum lycopersicum),eggplant (Solanum melongena),and pepper (Capsicum annuum)[J].Journal of Agricultural and Food Chemistry,2008,(06):1818-1828.doi:10.1021/jf073313n.
[9] Brunnert H J,Spener F,B(o)rchers T. PCR-ELISA for the CaMV-35S promoter as a screening method for genetically modified Roundup Ready soybeans[J].European Food Research and Technology,2001.366-371.doi:10.1007/s002170100371.
[10] Liu G,Su W,Xu Q. Liquid-phase hybridization based PCR-ELISA for detection of genetically modified organisms in food[J].Food Control,2004,(04):303-306.doi:10.1016/S0956-7135(03)00081-1.
[11] Jennings J C,Albee L D,Kolwyck D C. Attempts to detect transgenic and endogenous plant DNA and transgenic protein in muscle from broilers fed yield gard corn borer corn[J].Pour Sei,2003,(03):371-380.
[12] Gert Van Duijn,Ria Van Biert,Henri(A)ette BleekerMarcelis. Detection methods for genetically modified crops[J].Food Control,1999.375-378.
[13] Rogan G J,Dudin Y A,Lee T C. Immunodiagnostic methods for detection of 5-enolpyruvylshikimate-3-hosphate synthase in Roundup Ready soybeans[J].Food Control,1999,(6):407-414.doi:10.1016/S0956-7135(99)00083-3.
[14] 阚贵珍,喻德跃. 试纸条法和PCR法检测抗草甘膦转基因大豆的外源基因[J].中国油料作物学报,2005,(04):18-21.doi:10.3321/j.issn:1007-9084.2005.04.004.
[15] Notomi T,Okayama H,Masubuchi H. Loop-mediated isothermal amplification of DNA[J].Nucleic Acids Research,2000,(12):12.doi:10.1093/nar/28.12.e63.
[16] Yasuyoshi Mori,Kentaro Nagamine,Norihiro Tomita. Detection of loop-mediated Isothermal amplification reaction by turbidity derived from magnesium pyrophosphate formation[J].Biochemical and Biophysical Research Communications,2001,(01):150-154.
[17] Tomotada Iwamoto,Toshiaki Sonobe,Kozaburo Hayashi. Loop-mediated isothermal amplification for direct detection of Mycobacterium tuberculosis Complex,M.avium,and M.intracellulare in sputum samples[J].Journal of Clinical Microbiology,2003,(06):2616-2622.
[18] Akihiro Yoshida,Shiori Nagashima,Toshihim Ansai. Loop-mediated isothermal amplification method for rapid detection of the periodontopathic bacteria Porphyromonas gingivalis,Tannerella forsythia,and Treponema denticola[J].Journal of Clinical Microbiology,2005,(05):2418-2424.doi:10.1128/JCM.43.5.2418-2424.2005.
[19] Manmohan Parida,Kouhei Horioke,Hiroyuki Ishida. Rapid detection and differentiation of dengue virus serotypes by a real-time reverse transcription-loop-mediated isothermal amplification assay[J].Journal of Clinical Microbiology,2005,(06):2895-2903.
[20] Hiromi Ikadai,Hiroko Tanaka,Nona Shibahara. Molecular evidence of infections with babesia gibsoni parasites in Japan and evaluation of the diagnostic potential of a loop-mediated isothermal amplification method[J].Journal of Clinical Microbiology,2004,(06):2465-2469.
[21] Hiroki Hirayama,Soichi Kageyama,Satoru Moriyasu. Rapid sexing of bovine preimplantation embryos using loop-mediated isothermal amplification[J].THERIOGENOLOGY,2004.887-896.
[22] Shiro Fukuta,Yuko Mizukami,Akira Ishida. Real-time loop-mediated isothermal amplification for the CaMV-35S promoter as a screening method for genetically modified organisms[J].European Food Research and Technology,2004.496-500.
[23] 王永,兰青阔,赵新. 转基因作物外源转基因成分环介导等温扩增技术检测方法的建立及应用[J].中国农业科学,2009,(04):1473-1477.doi:10.3864/j.issn.0578-1752.2009.04.044.
[24] 梁蕊芳. 利用基因枪轰击法将NHX1、CBF耐逆相关基因导入高羊茅(Festuca arundinacea Schreb.)[D].内蒙古农业大学学报,2005.
[25] Murray M G,Thompson W F. Rapid isolation of high molecular weight DNA[J].Nucleic Acids Research,1980,(19):4321-4325.doi:10.1093/nar/8.19.4321.
[26] Nagamine K,Hase T,Notomi T. Accelerated reaction by loop-mediated isothermal amplification using loop primers[J].Molecular and Cellular Probes,2002,(03):223-229.doi:10.1006/mcpr.2002.0415.

Please choose a citation manager

Content to export