Identification of the Crude Drug Atractylodes Rhizome (Byaku-jutsu) and Atractylodes Lancea Rhizome (So-jutsu) Using Chloroplast TrnK Sequence as a Molecular Marker

• Published in: Biological & pharmaceutical bulletin 2000/05/01, Vol.23(5), pp.589-594
• Main Authors: MIZUKAMI, Hajime, OKABE, Yuka, KOHDA, Hiroshi, HIRAOKA, Noboru
• Format: Article
• Language: English
• Subjects: Asteraceae - chemistry; Asteraceae - genetics; Atractylodes Lancea Rhizome; Atractylodes Rhizome; Biological and medical sciences; chloroplast trnK; Chloroplasts - genetics; crude drug authentication; direct sequencing; DNA, Plant - analysis; Drugs, Chinese Herbal - analysis; General pharmacology; Genes, Plant; Genetic Markers; Medical sciences; Molecular Sequence Data; PCR-RFLP; Pharmacognosy. Homeopathy. Health food; Pharmacology. Drug treatments; Polymerase Chain Reaction; Polymorphism, Restriction Fragment Length; Sequence Analysis
• ISSN: 0918-6158; 1347-5215
• DOI: 10.1248/bpb.23.589

Novel methods for molecular authenticaiton of Atractylodes-derived crude drugs (Jutsu) were established based on PCR-restriction fragment length polymorphism (RFLP) and direct sequencing of chloroplast trnK. Two regions inside the chloroplast trnK were selected as molecular markers for identification and discrimination of Atractylodes Rhizome (Byaku-jutsu) and Atractylodes Lancea Rhizome (So-jutsu). The Region 1 fragment (260 bp) amplified from So-jutsu and Wa-byaku-jutsu (Atractylodes Rhizome derived from A. japonica) gave 2 bands of 180 bp and 80 bp on agarose gel electrophoresis after digestion with a restriction endonuclease HinfI, whereas the fragment amplified from Kara-byaku-jutsu (Atractylodes Rhizome derived from A. ovata) remained undigested, which allowed unambiguous identification of Kara-byaku-jutsu. By direct sequencing of Region 2 (436 bp) and comparison of the nucleotide sequence data sets we could not only discriminate Byaku-jutsu and So-jutsu but also identify the original plant species of each crude drug specimen. A simple and reliable protocol for rapid preparation of DNA suitable for PCR from as little as 1 mg of Atractylodes-derived crude drugs was also described.