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本研究基于DNA条形码技术对市售动物药材僵蚕进行分子鉴定研究,应用二维DNA条形码技术,建立跨平台信息交换的“互联网+”中药材物种鉴定体系。僵蚕药材同时包含动物家蚕和真菌白僵菌,采用植物基因组DNA提取试剂盒能成功扩增白僵菌ITS序列,未能扩增家蚕COI序列,而动物基因组DNA提取试剂盒提取总DNA均能成功扩增COI序列和ITS序列,并且ITS序列扩增、测序结果与植物基因组DNA提取试剂盒相一致。COI序列经鉴定为家蚕Bombyx mori Linnaeus,ITS序列经鉴定主要来源为白僵菌Beauveria bassiana(Bals.)Vuillant,少数为杂菌。基于ITS序列所构建的NJ树结果显示,白僵菌及其他致病菌株均可明显区分,且市售僵蚕药材混伪品含杂菌致病菌株。研究表明利用动物基因组DNA提取试剂盒可一次性将家蚕和白僵菌的DNA同时提取出来,有效解决同时含有动物和真菌类药材的基原鉴定。基于COI序列和ITS序列的DNA条形码分子技术能稳定、准确鉴别动物药材僵蚕,“互联网+”二维DNA条形码体系能有效监管药材流通领域的各个环节,有利于推动中药材市场的标准化和规范化。
In this study, based on DNA barcoding technology, commercial identification of silkworm (Bombyx mori L.) was carried out. Two-dimensional DNA barcoding technology was used to establish a cross-platform information exchange system for the identification of Chinese herbal medicines with “Internet +”. Bombyx mori contains both silkworm and fungus Beauveria bassiana, plant genomic DNA extraction kit can be successfully amplified Beauveria bassiana ITS sequence, failed to amplify the COI sequence of silkworm, and animal genomic DNA extraction kit can extract the total DNA can The COI sequence and ITS sequence were successfully amplified, and the ITS sequence was amplified. The sequencing result was consistent with the plant genomic DNA extraction kit. The COI sequence was identified as Bombyx mori Linnaeus, and the ITS sequence was identified as Beauveria bassiana (Bals.) Vuillant. The NJ tree based on ITS sequences showed that Beauveria bassiana and other pathogenic strains could be distinguished clearly. Studies have shown that the use of animal genomic DNA extraction kit can be a one-time silkworm and Beauveria bassiana DNA simultaneously extracted effectively solve both the animal and fungal medicinal base identification. The DNA barcode molecular technology based on COI sequence and ITS sequence can stably and accurately identify silkworm strains of animal medicine, and the “Internet + ” two-dimensional DNA barcode system can effectively supervise all aspects of medicine circulation and promote the standardization of Chinese herbal medicine market And standardization.