Binary transformation systems based on 'shooter' mutants of Agrobacterium tumefaciens: a simple, efficient and universal gene transfer technology that permits marker gene elimination.
A simple transformation procedure with a positive selection scheme using the expression of the isopentenyl transferase ( ipt) gene of transfer DNA (T-DNA) 'shooter' mutants of Agrobacterium tumefaciens was elaborated. After comparing several 'shooter' mutants we found that particular strains frequently produced phenotypically normal shoots after co-culturing with tobacco leaf explants. Shoots selected for normal phenotype showed apical dominance and could be rooted with the same efficiency as non-transformed shoots. When binary vectors were introduced into these strains, stably integrated binary vector T-DNA sequences were found in some regenerants, which were produced under non-selective conditions on growth-regulator-free medium. Such phenotypically normal transformants typically lacked a stably integrated ipt gene. Normal looking shoots could also be produced in tomato, muskmelon and sweet pepper.
Mihálka
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V
Agricultural Biotechnology Centre, Szent-Györgyi A. u. 4., 2100, Gödöllo, Hungary.
Balázs
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E
Nagy
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I
D016960:Agrobacterium tumefaciens / Q000235:genetics*; D002212:Capsicum / Q000235:genetics / Q000502:physiology; D002934:Cinnamates; D018548:Cotyledon / Q000235:genetics / Q000502:physiology; D028463:Cucumis melo / Q000235:genetics / Q000502:physiology; D046508:Culture Techniques / Q000379:methods; D004269:DNA, Bacterial / Q000235:genetics; D024881:Drug Resistance, Bacterial / Q000235:genetics; D018014:Gene Transfer Techniques; D005819:Genetic Markers / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D006921:Hygromycin B / Q000031:analogs & derivatives* / Q000494:pharmacology; D007612:Kanamycin / Q000494:pharmacology; D018551:Solanum lycopersicum / Q000235:genetics / Q000502:physiology; D009154:Mutation; D018515:Plant Leaves / Q000235:genetics / Q000502:physiology; D018520:Plant Shoots / Q000235:genetics / Q000502:physiology; D030821:Plants, Genetically Modified / Q000187:drug effects / Q000235:genetics* / Q000502:physiology; D012038:Regeneration; D014026:Nicotiana / Q000235:genetics / Q000502:physiology; D014170:Transformation, Genetic
D002934:Cinnamates; D004269:DNA, Bacterial; D005819:Genetic Markers; C036483:T-DNA; D006921:Hygromycin B; C026273:hygromycin A; D007612:Kanamycin