Identification of selected Thai medicinal plants by PCR-RFLP method

Abstract

Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method was applied to generate DNA marker for identification of the selected medicinal plants that closely resemble in morphological and histological characters. In the PCR-RFLP method, fragments of DNA are amplified from each sample with a set of specific primers and they are further examined for their nucleotide sequences to provide information for restriction enzyme cut site. Finally, DNA fragmentation pattern is generated by selected restriction enzymes. Murraya koenigii, Micromelum minutum and Clausena excavata were classified in family Rutaceae. Murraya koenigii which is reported to possess antioxidant, anticarcinogenic, anti-lipid peroxidetive, hypoglycemic and hypolipidemic properties [1] can be distinguished from M. minutum and C. excavata by the presence of 2 fragments of DNA after amplification of the internal transcribed spacer (ITS) of nuclear ribosomal RNA gene (rDNA) followed by digestion with restriction enzyme BsrBI. Amplification the ITS sequence and fragmentation with the restriction enzyme ApaI results in fingerprint pattern that is useful in discrimination of Zanthoxylum myriacanthum (Rutaceae) and Zanthoxylum limonella. Z. limonella and not Z. myriacanthum is reported to exhibit antimalarial and antituberculous activities [2]. PCR-RFLP based on maturase K gene sequence and PvuI digestion was proved to discriminate Kaempferia parviflora (Zingiberaceae) from Kaempferia galanga and Kaempferia marginata. K. parviflora has been reported to possess cytotoxicity against human cholangiocarcinoma cell lines [3].