ABSTRACT
The reporter genes GUS, NPTII and BAR, either separately or in combination, have been exploited to determine if DNA which can directly enter plants, circulate within the plant and enter nuclei, can also integrate into the genome in a manner which will permit gene expression. Feeding of either seed-derived or adventitious cut shoots of Solanum aviculare with the GUS gene followed by rooting of the shoots and growing on, resulted in all tissues of the plant showing GUS activity as detected cytochemically. Southern blot analysis of plants derived from the adventitious shoots confirmed the presence of the reporter gene in roots. Reporter gene expression was observed also in the F1 generation. If GUS and NPTII or GUS, NPTII and BAR were fed together, then in each case it was possible to have both expression and Southern blot confirmation of each of the genes. There was a relatively high rate of transformation of approximately 5% of the fed stems across all experiments conducted during the present study.
Subject(s)
DNA/metabolism , Plant Shoots/genetics , Solanum/genetics , Solanum/physiology , Transformation, Genetic , Anti-Bacterial Agents/metabolism , Biological Transport , DNA/administration & dosage , Drug Resistance , Genes, Reporter , Glucuronidase/genetics , Kanamycin/metabolism , Kanamycin Kinase/genetics , Plant Shoots/metabolism , Plants, Genetically Modified , Seeds/metabolismABSTRACT
A method for plant regeneration via organogenesis in pea (Pisum sativum) using nodal thin cell layer segments has been developed.From 10 to 12 days old sterile pea seedlings, nodal expiants were excised from which leaves and axillary buds were removed. Shoot regeneration was consistently obtained from liquid cultures where the expiants were floated on the medium. Shoots could be harvested after two weeks and thereafter up to ten weeks and no important effect of the cultivar (Bodil, Puget, Rondo and Trille) used could be observed as far as shooting capacity was concerned.Rooting frequency of the regenerated shoots was cultivar dependent. Plantlets were obtained within 7 weeks after expiant excision. Agrobacterium tumefaciens carrying a disarmed Tiplasmid and a binary vector containing the ß-glucuronidase reporter gene, were used in cocultivation experiments on pea nodal expiants in order to obtain transgenic shoots.
ABSTRACT
Induction of crown galls on 4-6 week old soybean (Glycine max (L.) Merr.) plants cultured in growth chambers was obtained with Agrobacterium tumefaciens strains C58, T37 and ACH5. The crown galls were isolated and cultured in vitro as sterile callus and liquid suspension cultures. Transformation was tested by opine tests and by molecular hybridization of restricted cell DNA with T-DNA fragments. Protoplasts were isolated from suspension cultures. Transformed protoplasts regenerate cell walls, divide and form calli without an exogenous supply of hormones.
