ABSTRACT
Objective To explore a new identification method for medicinal materials of Dysosma, and analyze the second internal transcribed spacer (ITS2) barcode sequences of Diphylleia sinensis and Dysosma versipellis, Sinopodophyllum hexandrum, Dysosma difformis and Dysosma pleiantha in five kinds of podophyllum. Methods The ITS2 of ribosomal DNA of medicinal materials of podophyllum was amplified and sequenced by bi-directional sequencing of PCR products. Sequence assembly and consensus sequence generation were performed by using CodonCode Aligner. Phylogenetic study was performed using software MEGA 5.1 in accordance with Kimura-2-parameter (K2P) model. Genetic distances were calculated and analyzed and the phylogenetic tree was constructed by using the neighbor-joining (NJ) method. Results There were significant differences among five kinds of Dysosma. Their maximum intraspecific genetic distance (K2P distance) was far lower than their minimum interspecific genetic distance with the other species. In the cluster dendrogram, all species showed monophyletic. Conclusion ITS2 sequence as DNA barcoding technique can be used to identify Chinese herbal materials of Dysosma.
ABSTRACT
Artemisia annua also known as Qinghao, is a kind of traditional Chinese medicine. Its active ingredient is artemisinin, a sesquiterpene lactone compound with a peroxy bridging group structure. A. annua is an effective antimalarial drug. Artemisinin, a secondary metabolite in A. annua, can be induced by many physical and chemical factors, such as salinity, moisture, light, and plant hormones. Temperature, as an important growth factor, also has a great influence on the synthesis of artemisinin. This article aims to study the effect of high temperature on inducing artemisinin biosynthesis in A. annua. The A. annua seedlings were placed at 25, 40 °C, and the samples were taken after 0, 3, 12 and 36 h. The content of artemisinin in each sample was determined by liquid chromatography-mass spectrometry. Total RNA was extracted from the samples, and then transcriptome sequencing and real-time fluorescence quantitative PCR were used to quantitatively analyze the expression of the key enzyme genes in artemisinin synthesis pathway and competition pathway. The results showed that artemisinin content was increased by 20%, 42% and 68% after 3, 12, 36 h of treatment at 40 °C. The expression levels of FDS, ALDH1, CYP71AV1 and ADS were up-regulated by 4.3, 3.3, 2.5, 1.9 times, and the expression levels of SQS and BPS were down-regulated by 37% and 90% respectively. In summary, high temperature can promote the biosynthesis of artemisinin by promoting the expression of synthetase genes in artemisinin synthesis pathway and inhibiting the expression of synthetase genes in artemisinin-competition pathway.