Study On The Induction Of Hairy Roots Of Sophora Tonkinensis Gapnep

The dried root and rhizome of Sophora tonkinensis Gapnep. is a commonly used traditional Chinese medicine with a long history of application. Matrine and oxymatrime are the chief active components of S. tonkinensis, and both with wide range of pharmacological actions, such as anticancer and antibiosis. The drug obtained from plant of S. tonkinensis cannot meet the medicinal and industrial used, so we try to find alternative resources or alternative way to produce raw materials for industrial extraction through Biotechnology. Provided alternative resource of S. tonkinensis through hairy root propagation is advantageous, such as genetic stability, fast-growing, strong synthesis ability of secondary metabolite and releasing some secondary metabolites to the culture medium, In this study, the aseptic seedlings of S. tonkinensis were infected to induce hairy roots by coculturing with Agrobacterium rhizogenes. The influence of culture method and growth regulator were evaluated to selected the most beneficial medium for hairy growth. To explore a new way to S. tonkinensis. cell and organ instead of raw material in the culture oriented S. tonkinensis., The main details of this research were as follow:(1) The A.rhizogenes R1601 was the most suitable strains and cotyledon was the most suitable explants for hairy roots inducing of S.tonkinensis induction. The highest induction rate was obtained from Ag.rhizogenes R1601, which was as high as 66.67%. And the hairy roots were generated from the wound of cotyledons by Ag.rhizogenes R1601 infected, which were stout, rapid growth.(2) Pre-culture S. tonkinensis axenic cotyledon for 2 days can improve the induction effect.(3)The concent of bacterial cultures with OD value of 0.6～1.0 showed high induction rate, and the highest induction rate was 72%, which obtained from OD value of 0.8.(4)The induction rate of the A.rhizogenes R1601 was increased with the time of infection increasing. But when the infection time was more than 16 min, the induction rate decreased.(5)Three days co-cultured after A. Rhizogenes R1601 infected of cotyledon ossess the highest onversion rate, which was as high as 72.00%. But the conversion rate decline when the co-culture over 3days.(6) The co-culture medium added with different concentrations of acetosyringone (AS) on hairy roots of S. tonkinensis was obvious. The highest induction rate was obtained at the AS concentration of 100μmol/L, which was as high as 66.67%.(7) Different concentrations of Cef possessed different antimicrobial effect on the receptors of cotyledon. High antibacterial effect was obtained with 300 and 500mg/L Cef. And considering the cost of saving,300mg/L Cef was the most suitable inhibitory concentrations for receptor.(8) 1/2MS culture medium was the best medium for the hairy roots growth, which have shown rapidly growth and no callus phenomenon.(9) IAA have significant correlation to the growth of hairy roots of radix sophorae tonkinensis, NAA and 6-BA on hairy roots of radix sophorae tonkinensis effect on growth was not significant and high concentration of NAA, hairy roots have tendency of callus. So the 1/2MS+0.6mg/1 IAA+0.5mg/1 NAA+0.1mg/1 6-BA is more suitable for hairy root culture S. tonkinensis.(10) By high performance liquid phase chromatography of radix S. tonkinensis hairy root is containing effective components, the total content of matrine and oxymatrine in the sample was 0.68%, but slightly lower than 0.7 per cent of the Pharmacopoeia, have medicinal value.