Tomato plant regeneration in genetic transformation

When tomato ZF seedlings grew 5-7 days after germination,cotyledons were cut into 0.5 cm× 0.5 cm discs and hypocotyls into 1 cm segments as explant.MS was used as basic medium,and Zeatin 1.0 mg/L and BA 1.0 mg/L were resperctively combined with IAA 0.05,0.2,0.5 and 1.0 mg/L as additions.It was determined that MS+BA 1.0 mg/L+IAA 0.2 mg/L was the best shooting medium.IAA 0.0 ,0.05,0.1 and 0.2 mg/L were respectively added into MS medium and rooting effects were tested.Results showed that the best rooting medium was MS+IAA 0.05 mg/L.Optimum concentration of Kan was 25 mg/L.Process of transformation culture was as followings:The discs and segments were pre cultured in shooting medium for 24 h (26 ℃,2 600 lx).Agrobacterium LBA4404 was cultured overnight in YEB and diluted 10-20 times with MS medium,in which explants were soaked for 5 minutes.Then the exptants were co cultured for 48 h (28 ℃,in the dark).After that,the explants were cultured in medium MS+BA 1.0 mg/L+IAA 0.2 mg/L+Kan 25 mg/L+Cef 200 mg/L for selection with transfer every two weeks.When shoots were 2 cm high,they were cut and transferred into the medium MS+IAA 0.05 mg/L+Kan 12.5 mg/L+Cef 100 mg/L for rooting.