PCR-Free Detection of Genetically Modified Organisms Using Magnetic Capture Technology and Fluorescence Cross-Correlation Spectroscopy 英文参考文献.docVIP

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PCR-Free Detection of Genetically Modified Organisms Using Magnetic Capture Technology and Fluorescence Cross-Correlation Spectroscopy 英文参考文献.doc

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PCR-Free Detection of Genetically Modified Organisms Using Magnetic Capture Technology and Fluorescence Cross-Correlation Spectroscopy 英文参考文献

PCR-FreeDetectionofGeneticallyModifiedOrganisms UsingMagneticCaptureTechnologyandFluorescence Cross-CorrelationSpectroscopy XiaomingZhou1,DaXing1*,YonghongTang1,WeiR.Chen2 1MOEKeyLaboratoryofLaserLifeScienceInstituteofLaserLifeScience,CollegeofBiophotonics,SouthChinaNormalUniversity,Guangzhou,China,2Departmentof EngineeringandPhysicsCollegeofMathematicsandScience,UniversityofCentralOklahoma,Edmond,Oklahoma,UnitedStatesofAmerica Abstract Thesafetyofgeneticallymodifiedorganisms(GMOs)hasattractedmuchattentionrecently.Polymerasechainreaction(PCR) amplification is a common method used in the identification of GMOs. However, a major disadvantage of PCR is the potential amplification of non-target DNA, causing false-positive identification. Thus, there remains a need for a simple, reliableandultrasensitivemethodtoidentifyandquantifyGMOincrops.Thisreportistointroduceamagneticbead-based PCR-free method for rapid detection of GMOs using dual-color fluorescence cross-correlation spectroscopy (FCCS). The cauliflower mosaic virus 35S (CaMV35S) promoter commonly used intransgenic products was targeted. CaMV35S target wascapturedbyabiotin-labelednucleicacidprobeandthenpurifiedusingstreptavidin-coatedmagneticbeadsthrough biotin-streptavidin linkage. The purified target DNA fragment was hybridized with two nucleic acid probes labeled respectivelybyRhodamineGreenandCy5dyes.Finally,FCCSwasusedtodetectandquantifythetargetDNAfragment through simultaneously detecting the fluorescence emissions from the two dyes. In our study, GMOs in genetically engineered soybeans and tomatoes were detected, using the magnetic bead-based PCR-free FCCS method. A detection limitof50pMGMOstargetwasachievedandPCR-freedetectionofGMOsfrom5mggenomicDNAwithmagneticcapture technology was accomplished. Also, the accuracy of GMO determination by the FCCS method is verified by spectrophotometry at 260nm using PCR amplified target DNA fragment from GM tomato. The new method is rapid andeffectiveasdemonstratedin