Identification of cardiotoxic ornamental plant species commonly found in Thailand by DNA analysis

Abstract

© 2017, International Society for Horticultural Science. All rights reserved. This research provided a forensic investigative tool for the detection and identification of cardiotoxic ornamental plant commonly found in Thailand. Species-specific primers were designed from polymorphisms in the ITS region of 6 cardiotoxic ornamental plant species, which are Adenium obesum, Calotropis gigantea, Cerbera odollam, Nerium oleander, Strophanthus gratus, and Thevetia peruviana. Sizes of expected PCR products from R_AO02/ITS3G, R_CG02/ITS3G, R_CO04/ITS5 R_NO12/ITS5, R_SG07/ITS3G and R_TP06/ITS3G are 358, 290, 282, 231, 226 and 165 bp, respectively. A positive test control primer pair was designed from a conserved region in the rbcL gene to confirm the validity of DNA sample for PCR-based analysis Under the optimal PCR condition, results showed that all primer pairs amplified PCR products of correct sizes. Amplification results were reproducible when tested against at least 10 individual plants of each species. Each of these primer pair did not PCR-amplify the other 5 non-target cardiotoxic plant species, 14 non-target plant species, nor human DNA. Correct test results were also obtained when tested with DNA mixture. For agarose gel-based sensitivity testing, the minimum DNA quantities that R_AO02/ITS3G, R_CG02/ITS3G, R_CO04/ITS5, R_NO12/ITS5, R_SG07/ITS3G and R_TP06/ITS3G can detect are 0.005, 0.001, 0.5, 0.01, 0.005 and 0.001 ng, respectively. Agarose gel-based analysis was possible for mock forensic-type specimens, such as dried leaf fragments and tea leaf infusions, but limit to specimens digested by artificial gastric juice.