Isolation and Culture of Protoplasts: Petunia

Binding, Horst; Krumbiegel-Schroeren, G.

doi:10.1016/b978-0-12-715001-7.50046-9

Cited by 3 publications

(1 citation statement)

References 27 publications

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“…Protoplasts were isolated from 5–6‐week‐old plants using an enzyme solution containing 0.4% cellulase ‘Onozuka’ R‐10, 0.4% macerozyme R‐10 (Yakult Pharmaceutical Co. Ltd, Japan), 0.06 m CaCl 2 and 0.375 m mannitol. Protoplasts were plated in culture medium V‐KM (Binding and Krumbiegel‐Schroeren, 1984) at a density of 10 5 cells ml −1 . After 3–5 days, when protoplasts started to divide, 50 µl Agrobacterium culture was added to the cell suspension.…”

Section: Methodsmentioning

confidence: 99%

Activation tagging identifies a gene fromPetunia hybridaresponsible for the production of active cytokinins in plants

Zubko¹,

et al. 2002

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SummaryCytokinins (CKs) are phytohormones that play an important role in plant growth and development. Although the ®rst naturally produced CK, zeatin, was isolated almost four decades ago, no endogenous gene has been shown to produce active CKs in planta. In an activation tagging experiment we have identi®ed a petunia line that showed CK-speci®c effects including enhanced shooting, reduced apical dominance and delayed senescence and¯owering. This phenotype correlated with the enhanced expression of a gene we labelled Sho (Shooting). Sho, which encodes a protein with homology to isopentenyl transferases (IPTs), also causes CK-speci®c effects when expressed in other plant species. In contrast to the ipt gene from Agrobacterium, which primarily increases zeatin levels, Sho expression in petunia and tobacco especially enhances the levels of certain N 6 -(D 2 -isopentenyl) adenosine (2iP) derivatives. Our data suggest that Sho encodes a plant enzyme whose activity is suf®cient to produce active CKs in plants.

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Section: Methodsmentioning

confidence: 99%

Activation tagging identifies a gene fromPetunia hybridaresponsible for the production of active cytokinins in plants

Zubko¹,

et al. 2002

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Sequential hormone requirement for growth and organogenesis of Petunia hybrida protoplasts-derived calli

et al. 1990

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Analysis of DNA polymerase activity in Petunia protoplasts treated with clastogenic agents

Benediktsson

Spampinato

Andreo

et al. 1994

Physiologia Plantarum

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Clastogenic agents, i.e. agents that can induce chromosome or DNA breakage, have been shown to enhance the rale of direct gene transfer to protoplasts. The effect was analysed at the enzymatic level using protoplast homogenates as well as intact protoplasts. For that purpose existing procedures were modified to enable measurement of DNA polymerase in vivo. In the system used, external DNA was able to enter the cells without the addition of membrane‐permeabilizing compounds. When comparing total DNA polymerase activity of protoplasts irradiated with X‐rays or UV‐light with that of untreated cells we did not observe significant differences. Incubation of protoplasts with high doses of bleomycin affected total DNA polymerase activity negatively. but dideoxythymidine triphosphate‐sensitive activity was not influenced. We conclude that the DNA strand‐breaks induced by low doses of X‐rays. UV‐light or bleomycin do not increase the total or the repair‐DNA polymerase activity and. therefore. that the increase in the transformation rates after DNA strand‐breaking is not preceded by enhanced DNA polymerase activity.

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Isolation and Culture of Protoplasts: Petunia

Cited by 3 publications

References 27 publications

Activation tagging identifies a gene fromPetunia hybridaresponsible for the production of active cytokinins in plants

Activation tagging identifies a gene fromPetunia hybridaresponsible for the production of active cytokinins in plants

Sequential hormone requirement for growth and organogenesis of Petunia hybrida protoplasts-derived calli

Analysis of DNA polymerase activity in Petunia protoplasts treated with clastogenic agents

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