Transgenic plantlets of 'Chancellor' grapevine (Vitis sp.) from biolistic transformation of embryogenic cell suspensions

Transgenic plantlets of 'Chancellor' grapevine (Vitis sp.) from biolistic transformation of embryogenic cell suspensions

Transgenic plantlets of 'Chancellor' grapevine (Vitis L. complex interspecific hybrid) were produced via biolistic transformation. Embryogenic cell suspensions were bombarded with 1 micrometer tungsten particles coated with pBI426 which encodes a fusion peptide between beta-glucuronidase (...

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Bibliographic Details
Published in:Plant cell reports Vol. 15; no. 5; pp. 311 - 316
Main Authors: Kikkert, J.R, Hebert-Soule, D, Wallace, P.G, Striem, M.J, Beisch, B.I
Format: Journal Article
Language:English
Published: Berlin Springer 1996
Subjects:
beta-glucuronidase
Biological and medical sciences
Biotechnology
cell suspension culture
culture media
dose response
embryo (plant)
enzyme activity
Fundamental and applied biological sciences. Psychology
gene expression
gene transfer
Genetic engineering
Genetic technics
genetic transformation
germination
kanamycin
methodology
Methods. Procedures. Technologies
neomycin
phosphotransferases (phosphomutases)
Transgenic animals and transgenic plants
Transgenic plants
Vitis
Cell culture
Microprojectile bombardment
Callus
Vegetals
β-Glucuronidase
Genetic transformation
Enzyme
Transferases
Vitis
Gene expression
Kanamycin kinase
Reporter gene
Vitidaceae
Dicotyledones
Glycosidases
Angiospermae
Liana
Hydrolases
Spermatophyta
Transgenic plant
O-Glycosidases
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Summary:Transgenic plantlets of 'Chancellor' grapevine (Vitis L. complex interspecific hybrid) were produced via biolistic transformation. Embryogenic cell suspensions were bombarded with 1 micrometer tungsten particles coated with pBI426 which encodes a fusion peptide between beta-glucuronidase (GUS) and neomycin phosphotransferase II (NPIII). 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 selecting 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 nonbombarded 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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ISSN:0721-7714
1432-203X
DOI:10.1007/bf00232362