Rare Medicinal Plants Changium Smyrnioides Germplasm Resource Protection And Rapid Propagation Research

Changium smyrnioides Wolff is a single specie of plants in Umbelliferae Changium in China,it distributs in the subtropical Yangtze River Valley,Zhejiang,Anhui,Jiangsu and Hubei provinces in east China’s areas.Changii cultivated in Jurong is popular for its strip and color.However,for human excavation and destruction of its habitat,its distribution area and individuals quantity are increasingly reducing.In recent years,the species tends to decline significantly,and Changium smyrnioides Wolff was included in three endangered plants in 1984.Since the Changium smyrnioides Wolff resources situation,in order to protect the existing resources,to protect Jiangsu Changium smyrnioides Wolff wild resources more effectively,to promote good preservation of germplasm resources,to provide high quality Changium smyrnioides Wolff clinical merchandise,and to provide new ideas and methods for protection and re-use the resource,We have carried out the following researches.（1） Changium smyrnioides Wolff resource survey and biological characteristics research.We have studied Changium smyrnioides Wolff in resource distribution,biological characteristics,resistance,trait identification,microscopical identification,finally we put forward several important protection measures to protect germplasm resources.（2） Changium smyrnioides Wolff ecological suitability research.Through ecological survey we find that its growth environment request is high and the suitable habitat is the evergreen and deciduous broad-leaved mixed forest in the limestone hillside,slope generally 10～40°.The analysis of soil structure,pH,organic matter contents showes that the suitable soil is yellow-brown and loam soil developed from calcareous limestone,pH 6.0～6.5,and needs of N,P,K fertilizer is different.The analysis of trace element shows that the accumulation of trace elements in medicine is closely related to the content of soil trace element.The average content of K is 28 times as much as Na,and content of Heavy metal all meet the requirements.Correlation Analysis shows that As has directly or indirectly Synergy with Essential elements of Cr、Fe absorption. （3）Based on the early study of Changium smyrnioides Wolff tissue culture,We have improved the rapid propagation methods further.The petiole and the leave were the best material explants in rapid propagation in all of the experiments.These explants were cultured on MS culture media by adding different portion of hormones at various cultural conditions.For the leaves,it is 2,4-D 4.0 mg·L^-1+Kt 0.4 mg·L^-1;Callus induction of petioles is little;for the adventitious buds,theyare6-BA2.0mg·L^-1+IBA0.4mg·L^-1 or 6-BA 3.0 mg·L^-1+IBA0.4 mg·L^-1;and for the roots,they are MS+IAA0.5 mg·L^-1 or MS+NAA0.2 mg·L^-1.Tissue culture of Changium smyrnioides Wolff makes its propagation rapid,its resources preserved,and its utilization last.（4） Changium smyrnioides Wolff preserving germplasm research.It was the first time to study on the cryopreservation technique of in vitro shoot-tips of Changium smyrnioides Wolff by vitrification and its plantet regeneration.Shoot-tips,1.5～2.0mm in size,excised from buds of in vitro shoots of Changium smyrnioides Wolff cold-hardened on the modified MS medium with nitrates reduced to half strength[MS（1/2N）]added with 0.3 mol·L^-1 sucrose at 6±1℃,that were precultured on[MS（1/2N）]medium with sucrose at concentration of 0.1,0.3 and 0.5 mg·L^-1 for 3 days.,were loaded with[MS（1/2N）]medium supplemented with 2 mg·L^-1 glycerol and 0.4 mg·L^-1 sucrose for 20 min at 22℃,and incubated in PVS₂ for 60 min at 0℃prior to direct plunge into liquid nitrogen.After rapid thawing for 2 min in water at 35±1℃,the shoot-tips were rinsed in[MS（1/2N）]medium supplemented with 1.2 mg·L^-1 sucrose for 20 min,plated on the regeneration medium supplemented with 2.0 mg·L^-1 6-BA,0.2 mg·L^-1 NAA,2.5 mg·L^-1 GA₃ sucrose and 8 g·L^-1 agar for lweek in dark prior to exposure to the light.The shoots formation was over 30%.（5） Changium smyrnioides Wolff chemical composition analysis.We adapt the "the Chinese Pharmacopoeia" method to compare the content of water soluble extracts and use HPLC to compare the fingerprints of Changium grown in differernts locations.