Abstract
Transgenic plantlets of ‘Chancellor’ grapevine (Vitis L. complex interspecific hybrid) were produced via biolistic transformation. Embryogenic cell suspensions were bombarded with 1 μm tungsten particles coated with pBI426 which encodes a fusion peptide between β-glucuronidase (GUS) and neomycin phosphotransferase II (NPTII). The fusion peptide is under the control of a double 35S Cauliflower Mosaic Virus promoter and a leader sequence from Alfalfa Mosaic Virus. The cells were placed on kanamycin-containing media (10, 25 or 50 mg/l) 2 d after bombardment. Activated charcoal reduced cell browning. Embryos were first observed on selective media 14–29 weeks after bombardment. More than 1600 clusters of embryos were germinated and/or assayed for GUS. Of 621 embryos assayed for GUS expression, 182 (29.3%) were positive. PCR confirmed the presence of the NPTII gene in all 5 GUS-positive and 2 GUS-negative (bombarded) embryos tested. In germination experiments, 15% of the embryo clusters produced at least one plant with normal shoot growth. Of 164 normal plants assayed for GUS expression, 37 (22.6%) were positive. The NPTII gene was amplified by PCR in 1 (of 1) GUS-positive and 4 (of 5) GUS-negative bombarded plants, but not in non-bombarded control plants. Southern blotting confirmed integration of the NPTII gene in all 3 of the GUS and PCR-NPTII positive plants tested. Biolistics is an efficient method for transformation of ‘Chancellor’ and should be applicable to other important grape cultivars.
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Abbreviations
- AC:
-
activated charcoal
- GUS:
-
β-glucuronidase
- 2,4-D:
-
2,4-dichlorophenoxyacetic acid
- BA:
-
6-benzylaminopurine
- NAA:
-
α-naphthalene acetic acid
- TDZ:
-
thidiazuron
- NPTII:
-
neomycin phosphotransferase II
- Km:
-
kanamycin
- MS:
-
Murashige and Skoog (1962) medium
- WPM:
-
Woody Plant Medium of Lloyd and McCown (1980)
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Communicated by G. C. Phillips
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Kikkert, J.R., Hébert-Soulé, D., Wallace, P.G. et al. Transgenic plantlets of ‘Chancellor’ grapevine (Vitis sp.) from biolistic transformation of embryogenic cell suspensions. Plant Cell Reports 15, 311–316 (1996). https://doi.org/10.1007/BF00232362
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DOI: https://doi.org/10.1007/BF00232362