Abstract
The objective of this study was to determine the efficacy of Agrobacterium tumefaciens in transforming spinach (Spinacia oleracea L.) callus. Callus was induced from leaf disks of `Baker' on Murashige and Skoog (MS) medium supplemented with 2 mg L-1 kinetin and 0.5 mg L-1 2,4-D. Callus was cut into 2-mm pieces, and 0.5 g of callus was placed in each 250-ml flask which contained 20 ml of MS liquid medium. The suspension cultures were inoculated with 100 μl of an overnight culture of A. tumefaciens harboring pMON 9749 (provided by S. Rogers, Monsanto Co., St. Louis), a plasmid cointegrated with kanamycin resistance and β -glucuronidase (GUS) genes. After coculturing for 2 days at 22C with shaking at 100 rpm, the medium was replaced with selection medium containing (in μg/ml) 75 kanamycin, 100 cefotaxime, and 200 carbenicillin and maintained for 3 weeks. Transient expression of GUS gene in transformed cells was detected with X-glu assay. This method resulted in a high level of transformation and provides the first report of transformation in spinach. This study was funded by a grant (92-B-32) from the Arkansas Science & Technology Authority.
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