The Transformation Of Tobacco By Ri Plasmid Of Agrobacterium Rhizogene And The Optimum Factors Affecting The Growth And Nicotine Biosynthesis Of Hairy Root

Factors affecting the transformation of tobacco by Ri plasmid of Agrobacterium rhizogene were studied and the optimum transformation conditions was defined. The structure of the hairy root tip and the dynamic characteristics of growth of a selected hairy root line were studied. The physical and chemical factors that affect the growth and nicotine biosynthesis of hairy root were investigated. The main results were as follows:1. Hairy roots were induced from tobacco leaves with Agrobacterium rhizogenes LBA1334 harboring a gropin type PRil855 and binary vector PI G121HM carrying a kanamycin resistant gene NPT II and GUS gene and the good hairy root clones were selected. The root with many root hairs and branches grew vigorously toward all the direction on kanamyain containing medium. The structure of the hairy root tip was not typical root cap. The culture system of hairy root could release nicotine into the medium and amount of nicotine was 96.3 % of the total content of nicotine produced by the culture system. Plantlet regerated from hairy root when cultured on MS medium without plant hormone.2. Factors affecting g the transformation of tobacco by Ri plasmid of Agrobacterium rhizogene were studied. Tobacco varieties, Agrobacterium rhizogene strains and basal medium influenced the efficiency of hairy root formation strikingly and the optimum transformation condition was defined.3. The growth of the hairy root and nicotine biosynthesis were influenced by various physical and chemical factors. It was found that the optimum pH value of medium was 6.0 and fast growth of the hairy root and maximal production of nicotine was observed in 1/2 medium and the presence of 20 g/L-30 g/L sucrose. The best volume of liquid medium for growth and nicotine synthesis of the hairy root was 2/5 of the volume of container at 110 rpm. IAA (1 mg/L) and LH (2 g/L) also enhanced the weight and nicotine content of the hairy root.4. Phytohormones affected obviously the growth and nicotine content of the hairy root. GA3 improved remarkably the growth speed of the hairy root but the nicotine synthesis was inhibited. IAA could raise the weight and the nicotine content of the hairy root when the concentration of IAA was 1 mg/L. The growth and nicotine content of the hairy root were not greatly promoted by addition of NAA.