| Peanut and sweet corn are both important and economic crops. They have abundant nourish value, good taste, most peoples are extremely like them, so their search value had been interested by scientific researchers and have become important food in biosphere. In recent years, a great deal of progress has been made in tissue culture and genetic transformation for peanut and maize. However, some problems have still been not solved such as low germination rates of somatic embryos, significant differences between geneotypes, abnormal embryos proportion and low transformation efficiency etc. So developing new regenerative system constantly, broadening the number of varieties of peanut and maize and trying to find out favorable transform conditions are a new trend presently in these two crops.The paper deals with the culture effect of the different explant from peanut seed and immature embryo of maize by organogenesis, et al. To test the regeneration efficiency of the explant getting ways of different genotype and the culture elements in peanut. Thinking that every analyses result systematical ,find it was found that leaflet is the excellent explant .So, the leaflets of peanut was genetically transformated through Agrobacterium-mediated method. A transformed plant was confirmed and the experiment factors influencing transformation efficiency were discussed in detail. All the results were summarized as follows:1. Immature embryos of sweet corn was used as explant on compound culture medium, 4 culture factors that affecting the maize regenerateds were investigated by experiment design and the best culture medium was found out . The results showed that AgNO3 was the key factor of typeâ…¡callus induction. 2, 4-D and L-pro take the second place, CH was the last. The optimal medium were N6(major)+B5(minor)+ Inositol (0.1g/l)+pro(500mg/l)+ 2,4-D(2mg/l) +AgNO3(10mg/l).2. Different explant from mature seed of peanut was used to test the plant regenerateion character esp.its'bud formation rate in vitro on different medium. Result indicated: all explant can induced callus and sprouting shoot ,but they had difference rate between explant: Epicoty>Blade>Hypocotyl. Medium's differences play a decisive role in induced callus and regeneration for different explant, and the same medium had different effect to different explant. It was found that in the experiment, pre-culture time had influence in buds induced and regeneration. Generally, in my result, short time pre-culture was better than its long time.3. The cultured effects of two kinds of leaflet from peanut were compared.The relationship was also discussed between the cultured element (genotype, medium, etc.)and cultured effects (dedifferentiation and redifferentiation).In the Pre-culture explant condition ,Variety SiLiHong six days explants on the 2nd medium differentiated 2.34 buds per callus, Variety GaiLiangHaiHua five days explants on the 1st medium regenerated 2.08 buds per callus.In the direct explants conditions, Variety GaiLiangHaiHua callus sprouting six buds and three from SiLiHong on the 1st medium . The cultured effect of direct explants was better than that of pre-cultured explants based on callus induction and differentiation. Variance analysis showed media had signioficant influence on the callus induction and differentiation, and genotypes were significant influence on callus induction, but not on sprouting rate.4. Leaflet from peanut varaiety SiLiHong and GaiLiangHaiHua was used as explant to culture on the different liquid culture medium. It was found that three basic mediums with various plant hormone combination can induce callus and sproute further. But by comparison the bud number on callus, the medium No.1 was better than the others. 3mg/l NAA was better than 1mg/lNAA while other hormone was same, Statistic analysis indicated that the genotype and medium had notable effects on calli induction and plant regeneration.5. The peanut genetic transformation was carried out by Agrobacterium-mediated approach on leaflet regeneration system. The factors affecting the transformation, such as the selective pressure of antibiotic, co-cultivation time, AS effect and the cultured effects of two kinds of leaflet were examined. The optimized procedure was as following: leaflet was co-cultivation with a vector containing Agrobacterium for 4 days on modified MS liquid medium (100 umol/l AS), 10 day's recovering cultivates, the explants were transferred to above selective media with 20mg/l Hy and 200mg/l Cef, 40-50 days later, the explants were transferred to above development media with 20mg/l Hy. 6 regeneration shoots were gained in first batch of material after 3 months culture. Out of these shoots, one was confirmed containing a characteristic 800bp DNA segment of the GUS by PCR analysis. The integration of foreign DNA into the peanut genome was confirmed in first step. |
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