Articles
Toward the development of Agrobacterium-mediated transformation system for the functional characterization of the genes involved in Prunus self-incompatibility
Article number
1408_16
Pages
121 – 127
Language
English
Abstract
Many Prunus species exhibit gametophytic self-incompatibility (GSI), of which pistil and pollen part specificities are controlled by S-ribonuclease (S-RNase) and S haplotype-specific F-box (SFB), respectively.
However, functional analysis of these genes remains to be performed because reproducible stable transformation system of Prunus has not been established for most Prunus species.
In this study, we tested a novel compound fipexide (FPX), and two genes ENHANCER OF SHOOT REGENERATION (ESR) and WUSCHEL (WUS), all reported to have promotive effects on regeneration, to develop Agrobacterium-mediated transformation system for the future use in functional analysis of Prunus self-incompatibility related genes.
When we tested the effect of FPX, callus formation but no adventitious bud formation was stimulated by 3, 10, 30, 100 and 300 µM FPX. In the transformation experiment using ESR and WUS with FPX, almost all explants formed callus in all conditions.
One each of ‘Benishuho’ explants regenerated shoots under the two different conditions.
One regeneration was observed with the explant transformed with ESR and cultured on medium containing 3 µM FPX for 2 days followed by 4.5 µM TDZ for 4 days, while the other regeneration was observed with an explant pre-cultured with 3 µM FPX and transformed with WUS, and cultured on medium containing 4.5 µM TDZ without FPX. Although the two regenerated shoots appeared to be escapes because they had no kanamycin resistance, no regeneration was obtained with control transformation.
Thus, it may be possible that regeneration was promoted by the effect of nearby transformed cells expressing ESR and WUS.
However, functional analysis of these genes remains to be performed because reproducible stable transformation system of Prunus has not been established for most Prunus species.
In this study, we tested a novel compound fipexide (FPX), and two genes ENHANCER OF SHOOT REGENERATION (ESR) and WUSCHEL (WUS), all reported to have promotive effects on regeneration, to develop Agrobacterium-mediated transformation system for the future use in functional analysis of Prunus self-incompatibility related genes.
When we tested the effect of FPX, callus formation but no adventitious bud formation was stimulated by 3, 10, 30, 100 and 300 µM FPX. In the transformation experiment using ESR and WUS with FPX, almost all explants formed callus in all conditions.
One each of ‘Benishuho’ explants regenerated shoots under the two different conditions.
One regeneration was observed with the explant transformed with ESR and cultured on medium containing 3 µM FPX for 2 days followed by 4.5 µM TDZ for 4 days, while the other regeneration was observed with an explant pre-cultured with 3 µM FPX and transformed with WUS, and cultured on medium containing 4.5 µM TDZ without FPX. Although the two regenerated shoots appeared to be escapes because they had no kanamycin resistance, no regeneration was obtained with control transformation.
Thus, it may be possible that regeneration was promoted by the effect of nearby transformed cells expressing ESR and WUS.
Publication
Authors
M. Kikunaga, R. Tao
Keywords
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