Study On DNA Barcoding Identification And Good Agricultural Practice Of Ligusticum Jeholense

Ligusticum jeholense Nakai et Kitag is one of the perennial herbs of Umbelliferae Ligusticum L. This plant is valued for its dried rhizomes which firstly occurred in Shennong’s Canon of Materia Medica. It can relieve common cold, parietal headache, rheumatism, joints associated with arthritis and scabies. Its ethanol extracts have stronger anti-inflammatory, sedative and hypnotic effets. Wild resources tend to extinction due to the great demand and intensive collection. Therefore, it is necessary to cultivate this medicinal plant. In this study, we accomplished the researchs on GAP technology of Ligusticum jeholense via identification, micro propagation, seed bionomics, cultivation and harvest time. The main results are as follows:1. We observed that the genetic diversity of Ligusticum jeholense could be well analyzed by DNA barcoding (NrITS, ITS2 and trnH-psbA) while eight DNA sequences (NrITS、rbcL、accD、 ITS2、trnH-psbA、ndhJ、rpoC1、rpoB) were analyzed.2. We developed an efficient micropropagation protocol for the multiplication of I. jeholense from immature embryo through adventitious bud proliferation. Immature embryos were cultured on Murashige and Skoog (MS) medium supplemented with 1.0 mg/L naphthaleneacetic acid (NAA),0.5 mg/L 6-benzylaminopurine (BAP) and 0.5 mg/L kinetin (Kn). About 40% of the explants germinated in vitro, and most of them produced the adventitious buds. In vitro-formed buds were successfully multiplied on a B5 medium containing 0.1 mg/L NAA,0.1 mg/L BAP and 1.0% agar, and a proliferation rate of 10.27-fold was steadily achieved. A bud clump containing three to five buds was successfully excised from the adventitious buds and transferred to the B5 medium supplemented with 0.6 mg/L NAA. About 92.17% of them successfully developed roots after 4 weeks; more than 80% of the rooted plantlets survived in a mixture of surface soil and vermiculite (v/v; 1:1;) in the greenhouse, and the direct transplanting to the field was also successfully established.3. We developed an efficient Standard Operation Procedure for the seed production of L. jeholense through the testing of germination temperature, germination substratum, kernel weight and comparison of light and dark. It’s appropriate for Ligusticum jeholense to germinate on paper in the 15 degrees Celsius. Both thousand kernel weight and hundred kernel weight could be used for seed weight test of Ligusticum jeholense. Light is not necessary for germination of Ligusticum jeholense.4. The density experiment showed that low-density (40 cm X 25 cm) and mid-density (40 cm X 30 cm) were higher than high-density (40 cm X 40 cm) on plant height, fresh weight of aerial part and root, stockpiled dry matter of aerial part and root, yield, content of ferulic acid and ligustilide. The fertilizer experiment showed that different N, P and K treatment have significant differences on fresh weight, stockpiled dry matter, yield, content of ferulic acid and ligustilide. Considering both yield and quality, using the lower N, P and K is better.5. The harvest time experiment showed that triennial was significantly higher than biennial on plant height, fresh weight of aerial part and root, stockpiled dry matter of aerial part and root, yield, content of ferulic acid and ligustilide. The fresh weight and content of ferulic acid and ligustilide of triennial root reached the max before harvest. The stockpiled dry matter of triennial and biennial reached the max at harvest.