The use of the PMI/mannose selection system to recover transgenic sweet orange plants (Citrus sinensis L. Osbeck).
A new method for obtaining transgenic sweet orange plants was developed in which positive selection (Positech) based on the Escherichia coli phosphomannose-isomerase (PMI) gene as the selectable marker gene and mannose as the selective agent was used. Epicotyl segments from in vitro-germinated plants of Valencia, Hamlin, Natal and Pera sweet oranges were inoculated with Agrobacterium tumefaciens EHA101-pNOV2116 and subsequently selected on medium supplemented with different concentrations of mannose or with a combination of mannose and sucrose as a carbon source. Genetic transformation was confirmed by PCR and Southern blot. The transgene expression was evaluated using a chlorophenol red assay and isoenzymes. The transformation efficiency rate ranged from 3% to 23.8%, depending on cultivar. This system provides an efficient manner for selecting transgenic sweet orange plants without using antibiotics or herbicides.
Boscariol
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RL
Laboratório de Biotecnologia Vegetal, Centro de Energia Nuclear na Agricultura, Universidade de São Paulo, Piracicaba 13 400-970 São Paulo, Brazil.
Almeida
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WA
Derbyshire
M T V C
MT
Mourão Filho
F A A
FA
Mendes
B M J
BM
D032084:Citrus sinensis / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development; D046508:Culture Techniques; D005632:Fructose / Q000494:pharmacology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D005819:Genetic Markers / Q000235:genetics; D008358:Mannose / Q000494:pharmacology*; D008359:Mannose-6-Phosphate Isomerase / Q000235:genetics* / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development; D013395:Sucrose / Q000494:pharmacology
D005819:Genetic Markers; D005632:Fructose; D013395:Sucrose; D008359:Mannose-6-Phosphate Isomerase; D008358:Mannose