Establishment of a real-time fluorescent recombinase polymerase amplification(RPA)for the Dendrobium officinale
Aims:This paper aims to establish a rapid identification method for Dendrobium officinale based on recombinase polymerase amplification(RPA).Methods:Assemblies of primers and probes targeting the rDNA internal transcribed spacer(ITS)region of D.officinale were designed and screened.The real-time RPA reaction condition was optimized,while the specificity,sensitivity and stability of the test were determined.Results:D.officinale could be specifically detected with 15 min at 39 ℃ using the established real-time fluorescent RPA method.No cross reactions were recorded with the DNA of D.monili forme,D.loddigesii,D.hercoglossum,D.devonianum and D.aduncum.The detection limit for the target DNA was 1.60×10-3 ng/μL with a high stability.Conclusions:The established real-time fluorescence RPA method is simple and rapid,and has high specificity,sensitivity and stability,which is applicable for rapid and accurate identification of D.officinale.
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