RESUMEN
Objective To design a pair of allele-specific diagnostic primer for authenticating Dendrobium huoshanense from other species of Dendrobium Sw.only by PCR.Methods Based on rDNA ITS sequences database of D.huoshanense and other species of Dendrobium Sw.,an allele-specific diagnostic primer pairs was designed.Diagnostic PCRs were performed using the primer with the total DNAs of 19(original) plants from Dendrobium Sw.as templates.The positive was the genuine.Results When the(annealing) temperature was raised to 60 ℃, the template DNA of D.huoshanense could be amplified whereas the diagnostic PCR of the rest species were all negative.Conclusion The diagnostic PCRs can authenticate D.huoshanense from other species of Dendrobium Sw.efficiently.Compared to the authentication method by sequencing DNA fragments and morphological traits etc.,the allele-specific diagnostic PCR is not only simple with time-saving but also practical and effective.
RESUMEN
Objective To develop a convenient and effective molecular authentication method for identifying Dendrobium nobile from other species of Dendrobium Sw.Methods Based on rDNA ITS sequences of D.nobile and other species of Dendrobium Sw.of Huangcaos and Fengdous,unusual sequence sites were confirmed and an allele-specific diagnostic primer pair was designed for D.nobile. Diagnostic PCR was performed using the primers with the total DNA of D.nobile and other species of Dendrobium Sw.as templates to establish the positive PCR condition for D.nobile.Results An about 300 bp DNA positive fragment was amplified from D.nobile with anneal temperature at 66 ℃ and anneal time at 40 s,whereas no any DNA fragment was amplified from the rest 39 species of Dendrobium Sw.Conclusion D.nobile could be effectively distinguished from other species of Dendrobium Sw.by diagnostic PCR with allele-specific diagnostic PCR,which is not only effective and specific,but also simple and timesaving,and has a comprehensive application prospects.