A method for determining the animals of origin in crude drugs using MinION, a compact next-generation sequencer

Abstract

We evaluated whether MinION, an inexpensive, portable sequencer, can be applied for identifying the animals of origin in crude drugs. Standard and nonstandard crude drugs with different species of origin were examined. In addition, the standards mixed with nonstandard samples were used. As a target gene, cytochrome c oxidase I was amplified and sequenced. The Fast mode results had a slightly lower match ratio than High-accuracy mode, but the animals of origin were correctly determined by BLAST for all samples. For antler velvet derived from Rangifer tarandus, even the sequences were aligned based on Cervus elaphus, the animal of origin was determined correctly. Minor contents could be detected from mixtures of two animals, if the mixtures contained at least 19:1 mtDNA when the coverage allele-fraction threshold was 0.05. By contrast, in Fast mode, two sequences could not be separated due to the low accuracy of the base-calling in each read. For field work, the species of origin of crude drugs could be identified, by only simple DNA extraction and library preparation. Therefore, MinION appears to be convenient tool for identifying the animal of origin in crude drugs.