title: Floral-dip transformation of flax (Linum usitatissimum) to generate transgenic progenies with a high transformation rate.
abstract: Agrobacterium-mediated plant transformation via floral-dip is a widely used technique in the field of plant transformation and has been reported to be successful for many plant species. However, flax (Linum usitatissimum) transformation by floral-dip has not been reported. The goal of this protocol is to establish that Agrobacterium and the floral-dip method can be used to generate transgenic flax. We show that this technique is simple, inexpensive, efficient, and more importantly, gives a higher transformation rate than the current available methods of flax transformation. In summary, inflorescences of flax were dipped in a solution of Agrobacterium carrying a binary vector plasmid (T-DNA fragment plus the Linum Insertion Sequence, LIS-1) for 1 - 2 min. The plants were laid flat on their side for 24 hr. Then, plants were maintained under normal growth conditions until the next treatment. The process of dipping was repeated 2 - 3 times, with approximately 10 - 14 day intervals between dipping. The T1 seeds were collected and germinated on soil. After approximately two weeks, treated progenies were tested by direct PCR; 2 - 3 leaves were used per plant plus the appropriate T-DNA primers. Positive transformants were selected and grown to maturity. The transformation rate was unexpectedly high, with 50 - 60% of the seeds from treated plants being positive transformants. This is a higher transformation rate than those reported for Arabidopsis thaliana and other plant species, using floral-dip transformation. It is also the highest, which has been reported so far, for flax transformation using other methods for transformation.
authors.affiliation: Department of Biology, Case Western Reserve University.
authors.forename: Nasmah K
authors.initials: NK
authors.lastname: Bastaki
authors.affiliation: Department of Biology, Case Western Reserve University; cac5@case.edu.
authors.forename: Christopher A
authors.initials: CA
authors.lastname: Cullis
mesh_terms: D060054:Agrobacterium / Q000235:genetics*; D017931:DNA Primers; D004269:DNA, Bacterial / Q000008:administration & dosage / Q000235:genetics*; D019597:Flax / Q000235:genetics* / Q000382:microbiology; D005822:Genetic Vectors / Q000235:genetics; D030821:Plants, Genetically Modified / Q000235:genetics* / Q000382:microbiology; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction; D012639:Seeds / Q000235:genetics; D014170:Transformation, Genetic
chemical_list: D017931:DNA Primers; D004269:DNA, Bacterial; C036483:T-DNA