| Maize is an kind of importment crop in the world as well as in China. In order to improve the quality of maize and enhance its production, it is important for us to apply the genetic engineering to transfer target gene to maize . It is applicable to use biolistic system to obtain transgenic corn plants. First of all ,if using biolistic system,it is to erect regenable receptor system. In the paper, the method to establish a better receptor system of maize inbred line for biolistic bombarderment was studied.Many factors which affect callus induction,subculture and ^differentiation were studied. On the basis of this work,the receptor system were successfully built up by using immature embryo of maize inbred line 7922 as model.Subsequently,immature embryos of inbred line Pl,P2,H99,Mol7 also were successfully inoculated, and type-II callus were obtainedThe results of the experiment of induction of callus showed that MB medium is the better medium than other media tested,such as 8114,MS,N6,NMB,AG.The frequency of induction of callus was higher and the callus which inducted from MB was better also. Suitable 4"C treatment in a refrigerator(<7 days) prior to immature embryo dissection was an efficient way to improve the frequency of inocution. AgNOsdid not contribute to callus iniation in the study 20g/l sucrose plus 10g/l glucose was suitable carbon source for iniation medium. Immature embryos placed embryo-axis dow(scutellum side up) on callus initiation medium was better. Othenvise,the frequency of iniation was very low.It is essential for maize callus to transfer from Type-I to Type-II to build up receptor system which is suitable to transfer target gene with biolistics. DMSmedium could significantly expedise the process. The frequency of transformation of callus on DMS medium was about 48.28 per cent while those of MS,N6 were respectively around 15.63 per cent and 12.12 per cent.Factors influncing the subculture of Type-II callus ,such as kinds of media,concenttrtion of mannitol,carbon source,AgNC>3 and active charcoal were observed. The results showed that N6 and NMBwere two kinds of optimal media for subculture oframaize calus. The callus growed on them was pale yellow or yellow,compact,and friable. Five per cent mannitol which was added to subculture medium was benefitial to enhance the state of callus, at the same time,it could not influence speed of growth. Optimal carbon source in the subculture medium was 50g/l sucrose.At this concentration,sucrose not only provided enough energenic matter to callus,but also maintain a better osmotic condition of medium The results also showed that most callus seemed difficult to be maintained on medium added some active charcoal.Just lg/1 AC could make callus diffrentiation.AgNOs actually did the same effect on the process of subculture.After these factors were analyzed,optimal contition of subculture was established.Many factors affecting regeneration of Type-II callus, including kinds of medium, sorts of plant growth regulation matters and their concentration were studied.Optimal condition for rediffrentiation of Type-II callus was obtained.MS medium plus l.Omg/1 6-BA and 0.4mg/l IB A or N6 medium plus l.Omg/1 6-BA were two better media, espiecially the former The frequency of rediffrentiation was 105.26 per cent.The experiment of basic endurance of callus to Basta was done The result showed optimal concentration for screening after bombarderment was 6mg/l which could evidently blocked the growth of non-thansformation callus.In view of obtaining efficient transformation condition to maize cells by using biolistic system,factors influencing the frequency of transformation, rupture pressure and target distance, bombarderment times,concentration of Au powder and DNA,osmotic treatment for callus,and so on were researched when the Type-II callus of maize inbred line 7922 was taken as receptor system of bombardment.The results showed that optimal biolistic rupture pressureand target distance was HOOpsi X 9cm,optimal bombardment times was twice,suitable co... |
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