ISHS Acta Horticulturae 734: VIII International Symposium on Plum and Prune Genetics, Breeding and Pomology
PRODUCTION OF TRANSGENIC PLUM PLANTS FROM VEGETATIVE TISSUES BY MEANS OF POSITIVE SELECTION |
| Authors: | R.V. Mikhailov, S.A. Muratova, S.V. Dolgov |
| Keywords: | GFP, pmi, regeneration, transformation, plum |
| Abstract:
The most limiting factor in the plum genetic transformation is the very pour regeneration efficiency of plum somatic tissues. For this reason, there are no reports on successful transformation of commercial plum varieties. The efficient protocol for plum plant regeneration and genetic transformation has been developed. An efficient protocol for adventitious shoot regeneration from plum leaf tissues was established by manipulating phytohormone types and concentrations, explant pretreatment and physiological age of explants. This protocol resulted in a regeneration rate that was sufficiently high (80%) to be useful for future Agrobacterium-mediated transformation work. Leaves of the plum cultivar ´Startovaya´ produced in vitro were infected with A. tumefaciens strain CBE 21 containing a binary vector pNOV35S-GFP. It contains the synthetic codon optimized red-shifted gfp gene driven by the 35S promoter and pmi gene (phosphomannose isomerase gene derived from E. coli) under the CMPS promoter from Cestrium Yellow Leaf Curling Virus. Plant cells with stably incorporated T-DNA exhibited cell division and developed transgenic calli, followed by formation of transgenic shoots. Stable integration of the pmi gene into the genome of 'Startovaya' plum was confirmed by PCR analysis. Expression of gfp gene was detected by fluorescent microscopy. | |
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