Agrobacterium-mediated transformation of commercial mints |
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Authors: | C Berry J M Van Eck S L Kitto A Smigocki |
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Institution: | (1) Delaware Agricultural Experiment Station, Department of Plant and Soil Sciences, College of Agricultural Sciences, University of Delaware, 19717-1303 Newark, DE;(2) Plant Molecular Biology Lab, Department of Agriculture, Agricultural Research Service, 20705 Beltsville, MD, U.S.A |
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Abstract: | Commercial peppermint (P) (Mentha × piperita L. ev. Black Mitcham), native spearmint (NS) (M. spicata L.) and Scotch spearmint (SS) (M. × gracillis Sole cv Baker) petioles and orange mint (OM) (M. citrata Ehrh.) leaf disks were cocultivated with a number of Agrobacterium tumefaciens strains. P, SS and OM initiated tumor-like callus tissue on growth regulator-free MS medium after cocultivation with strain A281, a hypervirulent agropine strain containing Ti plasmid pTiBo542. Callus did not initiate from explants cocultivated with strain C58, a virulent nopaline strain; with A 136, a plasmidless strain, or from uninoculated controls. A281-derived callus was maintained on growth regulator-free medium in the absence of antibiotics for up to two years with no bacterial outgrowth. No shoots regenerated from any of the tumors on regeneration medium. Five of seven OM callus lines assayed gave a positive signal for agropine. DNA extracted from OM tumor tissue hybridized to a DNA probe specific to the T-DNA region of pTi plasmid. Genomic Southern analysis of DNA from tumors of P and SS indicated that one to a few copies of the T-DNA integrated into the mint chromosomes. PCR amplification of genomic DNA with primers specific for one of the T-DNA encoded genes yielded fragments that, when analyzed by restriction enzyme mapping and on Southern blots, corresponded to the cytokinin biosynthesis gene ipt. These results demonstrate transformation of three species of mint and the potential for using A. tumefaciens to transfer economically important genes into commercial mint cultivars.Abbreviations BA
benzyladenine
- CW
coconut water
- Cef
cefotaxime
- P
peppermint
- SS
scotch spearmint
- NS
native spearmint
- OM
orange mint
- BM
basal medium
- MS
Murashige and Skoog (1962)
- PAR
photosynthetically active radiation
- CTAB
hexadecylatrimethylammonium bromide
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ipt
isopentenyl transferase
Received for publication 1994. Published as Miscellaneous Paper No. 1482 of the Delaware Agricultural Experiment Station. Contribution No. 317 of the Department of Plant and Soil Sciences. Mention of trade names in this publication does not imply endorsement by the Delaware Agricultural Experiment Station of products named, nor criticism of similar ones not named. |
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Keywords: | Callus DNA slot blot hybridization Mentha polymerase chain reaction Southern analysis tissue culture |
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