Rapid identification of Colchicum autumnale based on loop-mediated isothermal amplification (LAMP) assay

• Published in: Forensic toxicology 2021-01, Vol.39 (1), p.259-265
• Main Authors: Misawa, Sae, Natsuhara, Daigo, Kiba, Yuka, Yamamuro, Tadashi, Suzuki, Ryuichiro, Shibata, Takayuki, Kitamura, Masashi
• Format: Article
• Language: English
• Subjects: Forensic Medicine; Forensic Science; Medical Law; Medicinal Chemistry; Medicine; Medicine & Public Health; Pharmacology/Toxicology; Short Communication
• ISSN: 1860-8965; 1860-8973
• DOI: 10.1007/s11419-020-00557-4

Purpose Colchicum autumnale (Colchicaceae) exhibits toxicity, and severe poisoning cases due to ingestion of this plant have been reported in Japan. Identifying the cause of poisoning is important for emergency medical treatment, and a rapid and simple detection technique is required for the identification of the cause of poisoning. In the present study, we developed a rapid and simple method for the detection of C. autumnale based on a loop-mediated isothermal amplification (LAMP) assay, which is 100- to 1000-fold higher than the conventional polymerase chain reaction method. Methods Specific LAMP primers for C. autumnale were designed based on the trn L -trn F intergenic spacer region. Using the LAMP primers, the LAMP assay was performed at an isothermal reaction temperature of 63 ℃. Results The LAMP reaction was shown to be specific and highly sensitive to C. autumnale , given that the assay can be used for 10 pg of purified DNA. Using a simple protocol for on-site detection, the entire procedure from pretreatment to evaluation required approximately 1 h. Moreover, the LAMP method using a microfluidic device detected multiple genes, including C. autumnale -specific DNA regions, at a time. Conclusions The currently proposed protocol exhibits good potential as a screening method for C. autumnale poisoning in emergency medical care.