title: Co-transformation of grapevine somatic embryos to produce transgenic plants free of marker genes.
abstract: A cotransformation system using somatic embryos was developed to produce grapevines free of selectable marker genes. This was achieved by transforming Vitis vinifera L. "Thompson Seedless" somatic embryos with a mixture of two Agrobacterium strains. The first strain contained a binary plasmid with an egfp gene of interest between the T-DNA borders. The second strain harbored the neomycin phosphotransferase (nptII) gene for positive selection and the cytosine deaminase (codA) gene for negative selection, linked together by a bidirectional dual promoter complex. Our technique included a short positive selection phase of cotransformed somatic embryos on liquid medium containing 100 mg/L kanamycin before subjecting cultures to prolonged negative selection on medium containing 250 mg/L 5-fluorocytosine.
authors.affiliation: Citrus Research and Education Center, University of Florida/IFAS, Lake Alfred, FL, USA.
authors.forename: Manjul
authors.initials: M
authors.lastname: Dutt
authors.forename: Zhijian T
authors.initials: ZT
authors.lastname: Li
authors.forename: Sadanand A
authors.initials: SA
authors.lastname: Dhekney
authors.forename: Dennis J
authors.initials: DJ
authors.lastname: Gray
mesh_terms: D016960:Agrobacterium tumefaciens / Q000235:genetics; D043525:Cytosine Deaminase / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D018014:Gene Transfer Techniques; D005819:Genetic Markers; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics; D019868:Kanamycin Kinase / Q000235:genetics; D030821:Plants, Genetically Modified; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic; D012639:Seeds / Q000235:genetics; D046509:Tissue Culture Techniques; D014170:Transformation, Genetic; D027843:Vitis / Q000196:embryology / Q000235:genetics* / Q000254:growth & development
chemical_list: D004269:DNA, Bacterial; D005819:Genetic Markers; C036483:T-DNA; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D019868:Kanamycin Kinase; D043525:Cytosine Deaminase