农杆菌介导剑叶文心兰遗传转化体系中抗生素的筛选OA

To optimize the efficient and stable genetic transformation system of Oncidium ensifolium,in vitro leaves were used as explants in this study.The effects of 3 types of selective antibiotics on callus and the efficacy of 2 types of bacteriostatic antibiotics were investigated via concentration gradient assays and mathematical/statistical analyses.Results showed that,as revealed by Duncan's multiple range test and analysis of variance(ANOVA),0.18 mg/L Basta,9 mg/L hygromycin(Hyg),and 100 mg/L kanamycin(Kana)completely inhibited adventitious bud differentiation in leaves(P＜0.01),and thus were identified as the selection pressure for resistant buds.The half-lethal concentrations(LC50)for subcultured seedlings were 0.12 mg/L Basta,5 mg/L Hyg,and 40 mg/L Kana(P＜0.05),which were designated as the screening concentrations for resistant seedlings.O.ensifolium exhibited antibiotic sensitivity in the order of Basta＞Hyg＞Kana(all P＜0.01),indicating that the bar gene is more suitable as a selectable marker.Additionally,100 mg/L cefotaxime(Cef)effectively inhibited Agrobacterium proliferation while maintaining a high leaf survival rate.This study provides critical parameters for optimizing the genetic transformation system and screening transgenic plants in Tolumnia.