Study On Evaluation Of Drought Stress Tolerance In 6 Petunia Cultivars And Transformation Of P_SARK-IPT Gene Into Petunia

Petunia is one of the common garden ornamental flowers.It has important value to screen strong drought resistance cultivars,under the background of the demand of "Conservation oriented landscape" construction in our country.Based on six different cultivars of Petunia,germination rate,germination potential,germination index,the average days of germination were measured under 4 kinds of PEG-6000 concentration gradient.13 indexes which were related to plant morphology,photo synthetic characteristics and physiological changes were measured,under 4 field RWC gradients.Then,a comprehensive evaluation and analysis was made to determine the drought resistance of Petunia’s seed and plantlet by using the membership function formula.And the P_SARK-IPTgene was transferred into the cultivar with poorest drought resistance,to improve its drought resistance.The results are as follows:（1）Under the drought stress circumstance,the germination rate,germination potential,germination index of Petunia’s seeds of 6 cultivars declined gradually and the average days of germination their gradually rose with the increase of concentration of PEG-6000.The experiment showed that each cultivar seed’s germination were subject to varying degrees of inhibition,with "Merlin" in the lead.The experiment of drought stress in plants showed that the plant flower diameter,plant height and the flowering rate decreased gradually with the decrease of soil moisture content.The net photosynthetic rate and the transpiration rate decreased by degrees,the WUE rose firstly and then reduced,the relative water content and chlorophyll content of leaf gradually reduced.The activity of SOD,POD and CAT and the content of MDA,soluble protein content showed a trend of increase.The range of variation of different cultivars is different.The period of seed germination and plant stage drought resistance of the order was obtained by using the membership function formula,and the order is same,from strong to weak: ‘Linbo’ > ‘Bold’ > ‘Ultra’ > ‘Sonja’ > ‘Eagle’ > ‘Merlin’.In conclusion,the drought-resistantability between seed germination and seedling stage is related.The cultivar has strong drought resistance during seed germination,and the seedling stage is alike.（2）The regeneration systems of ‘Merlin’was established.The result showed that MS +2.0mg·L-16-BA +0.3mg·L-1NAA was the optimum medium for callus and bud differentiation,the callus induction rate and adventitious bud induction rate were 100%,7.31 respectively.The optimal hormone concentration of inducing stem proliferation was 0.5 mg·L-1ZT,and the number of prociferation was 6.48.The best optimum rooting hormone concentration is 0.05mg·L-1IBA,the rate of rooting was 95.34%,and the number of rooting was 7.95.The survival rateof was 96.67% when the seedless seedlings were transplanted into the soil.（3）An efficient genetic transformation system was established.To improve drought resistance,transfer P_SARK-IPT gene into petunia.Using Hey as the selection marker in genetic transformation,experimental results showed that 2mg·L-1Hey was suitable for differentiation selection,and 4mg·L-1Hey was appropriate for rooting.The gene was transformated into Petunia by the genetic transformation system 500mg·L-1Cef as bacteriostatic and pre-culture time for 2d,infection time for 3min,co-cultivation time for 36 h,OD600=0.5,and obtained 15 transgenic resistant plants.PCR analysis suggested that the foreign gene had been transferred into 12 plants.Selected 8 transgenic resistant plants for RT-PCR detection found that 7 plants were positive plants at the reverse transcription level.