Abstract:The pungent condiments in the Japanese dishes are mainly made of Eutrema wasabia, Armorica Rusticana and Mustard, which belong to Brassicaceae plants. This study used the internal transcribed spacer II (ITS2) technology to quickly and accurately identify wasabi, mustard and Armorica Rusticana, providing a reference for the technology used in the quality testing of Pungent products. In this study, three plant materials (Eutrema wasabia, Armorica Rusticana and white mustard) were used as experimental materials to extract the genomic DNA, and the ITS2 sequence was obtained by PCR amplification using the universal primer ITS2_S2. After sequencing, bioinformatics analysis was performed for species identification. MEGA7.0 (Molecular Evolutionary Genetics Analysis 7.0) analysis of ITS2 sequence showed that the K2P genetic distance (Kimura 2-parameter) of wasabi, white mustard and horseradish is between 0.88 and 0.171, which are all more than 0.01, and there are 36 interspecies variation sites, which preliminarily concluded that the we can distinguish the three species of wasabi, mustard and horseradish with ITS2 sequence. In addition, ITS2 sequences of eutrema wasabi and its close relatives, such as eutrema edwardsii, Armorica Rusticana and mustard, were obtained from GenBank. MEGA7.0 was used to analyze interspecies sequence, calculate K2P and build evolutionary tree by Neighbor-joining (NJ). Eutrema wasabi and eutrema edwardsii are gathered into a large branch, in the meantime, brown mustard, white mustard, and yellow mustard are gathered into a large branch and Armorica Rusticana alone. ITS2 sequence analysis of eutrema wasabi and its close relatives, such as eutrema edwardsii, Armorica Rusticana, and mustard showed that the genetic distance of K2P between species was between 0.030 and 0.105, which are all more than 0.01. This study shows that ITS2 technology can be used to identify relative species such as wasabi, Armorica Rusticana and mustard. This technology can be used to identify and quantify the specific raw material ingredients of pungent condiments, providing a scientific basis for food quality control and food safety.
