Study on identification of Chinese herbal medicine by ITS rDNA sequences

• Main Authors: Tzung-ting Shie, 謝宗廷
• Other Authors: Tang-yao Hong
• Format: Others
• Language: zh-TW
• Published: 2013
• Online Access: http://ndltd.ncl.edu.tw/handle/11112609956966389196

碩士 === 大仁科技大學 === 生物科技研究所 === 101 === The ninety percent of Chinese herbal medicine in Taiwan has been imported from China. It is said that the quality of Chinese herbal medicine imported is not uniform with overdose heavy metal and pesticide residue. Therefore, the control of quality in Chinese herbal medicine is a significant issue for compatriots’ health. First of all, the Chinese herbal medicine strain is confirmed from the sources in the control of quality. The traditional identification in Chinese herbal medicine gives priority to appearances, tissue slice, and thin layer chromatography. In recent years, some scholars have assisted in the origin identification of Chinese herbal medicine by means of the molecular biology method. Analysis in this thesis, compared with the nucleotide sequences of the internal transcribed spacers (ITS) of the ribosomal, among several kinds of Chinese herbal medicine produced in Taiwan, is to be aware if this sequence can be taken as the auxiliary tool for the origin identification. The layouts in different Chinese herbal medicine primers, during the experiment, are to compare with the ITS sequence from the different species of the same genus in Chinese herbal medicine NCBI GenBank, including Lonicera spp., Salvia miltiorrhiza, Andrographis paniculata, Dioscorea spp., Taraxacum spp., and etc. After proceeding as the multiple sequences alignment in utilization of software, we design diverse pairs of conserved primers; then make good use of liquid nitrogen to grind Chinese herbal medicine for its chromosome DNA. To amplify its sequence in ITS rDNA by Polymerase Chain Reaction (PCR) method, to connect the PCR product and the TA cloning vector, to deliver clone to Biotechnology company for the sequencing, as well as to make sequence alignment from the sequence we got by NCBI GenBank are required. The identity of sequence in Salvia miltiorrhiza, Dioscorea spp., and Taraxacum spp. between grown in Taiwan and gotten in GenBank reaches more than ninety-nine percent. Again, the diversity among other different species of the same genus is much higher. It is a proof that these three sorts of Chinese herbal medicine can be made into the sequence identification by ITS rDNA. The identity of sequences in Lonicera spp. collected from distinct habitants in Taiwan reaches ninety-nine percent; however, the identity of Lonicera spp. in China or USA GenBank reaches merely eighty-three percent. Two possibilities are as follows: one, Lonicera spp. grown in Taiwan is endemic species, and the other, the amplifying sequences in Lonicera spp, is another set of ITS. Therefore, the sequence in GenBank is much far from the one grown in Taiwan. Owing to the result from those experiments, it is known that the ITS sequences analysis method cannot be identified effectively as five kinds of Chinese herbal medicine origin but possibly as the auxiliary tool. Keywords: Chinese herbal medicine, internal transcribed spacers, identification