Tobacco Cells And Establishment Of Hairy Root Culture System And Nicotine Bio-synthesis Of Research

Nicotine is one of the important botanic pesticidal substance which has been accepted in the world. Cell and hairy roots of Nicotiana tabacum (NicotianarusticaL.) cultures for producing nicotine were studied in this paper, with the aim of providing basic data and theories for producing nicotine and other botanic pesticidal substance. Sterile explants were got from three different ways , sterile seedling, asexual generation of seedling and regenerration of embryonic callus .Explants were cultured on 1S NT, B5, F basic medium with 2, 4 KT. NAA 6A at different concentration to induce callus .We got the most suitable callus and culture medium ( NT--2 4D2mgLTO. 25mg L) for establishing of cell suspension. The cell suspension grew fast. In the coulture of hypocotyl, stem . leaf of tahacum and A4 stain of agrohacterium i-h izogenes , the hairy roots of tahacum tansformed by A stain were induced. Hiary roots were identified by their hioharacter and agrohacterium rhizogenes A1 stain was completely eradicated by different endurance of antibiotic and temperature between agi-ohacvei-iwn -hizogenes and plant tissue .We got hairy root stain :3,4.47 in MS hormone free medium. Nicotine content of cell suspension and hairy roots were detected by the method of steam distillationV spectrophotometer scan which was prescripted in IS0288l. Nicotine production mediumMS4 wasfoundhvexperimentation.Cell suspension of tahacum cultured in medium MS, produced nicotine . The maximal nicotine content was 0.358% in the compound of cell and medium , 0. 23. in the liquid medium , 2.05% in dry weight of cell at 22 day of production culture. The hairy toot stain 4 was selected as topmost nicotine production stain among all the hair root stains in this experimentation. To improve nicotine production of hair?roots we got Improved Medium MS,, The hairy root stain 4 cultured in this medium yield the nicotine content at 0. 425% in the liquid medium, 4.050 in dry weight of hairy roots. . highly yielded hormonendependent callus line which nicotine yield at 0.332% in fresh weight. 4. 16% in dry weight of the caulls was also obtained. From our study ,we drew the conclusion that the form of nitrogen nutrition, the supply of some amino acid .the regulation of cell differentiation by hormone and the sucrose concentration were the crucial factor for the nicotine bio? production of cell culture in vitro and that the nicotine got from tahacum cell and hairy root culture in vitro is no different from standard nicotine as pesticide. The conclusion can he pro\red that nicotine producing through hioengineering way is feasible.