Search

Agrobacterium tumefaciens was used to transform fiber flax with the pBITUBA8 plasmid carrying the mutant alpha-tubulin gene imparting resistance to dinitroaniline herbicides and the nptII selective marker gene imparting resistance to canamycin. The transformants were selected in parallel on media containing canamycin and trifluralin (a dinitroaniline herbicide). The transgenic nature of the resultant regenerants resistant to dinitroaniline herbicides was confirmed by means of Southern blotting and polymerase chain reaction (PCR) analysis using specific probes for the ntpll gene and the gene of alpha-tubulin.

Biolistic transformation of soybean by construction carrying a mutant alpha-tubulin gene as selective marker gene conferring resistance to dinitroaniline herbicides, and by additional construction pAHTUBI carrying a full-length barley beta-tubulin gene for correct co-expression of exogeneous tubulin in cells of transgenic soybean lines has been realized. It was established that 10 microM trifluralin is the most optimal selective concentration to pick up transformed soybean lines. Transgenic nature of offained regenerants was confirmed by Southern blotting hybridization using specific probe to selective alpha-tubulin gene.

Tubulins, as major components involved in the organization of microtubules, play an important role in plant development. We describe here the expression profiles of all known alpha-tubulin (TUA), beta-tubulin (TUB) and gamma-tubulin (TUG) genes of barley (Hordeum vulgare), involving eight newly identified TUB sequences, five established TUA genes and one TUG gene. Macroarray and Northern blot-based expression patterns in the pericarp, endosperm and embryo were obtained over the course of the development of the grain between anthesis and maturation. These revealed that the various tubulin genes differed in their levels of expression, and to some extent were tissue specific. Two expression peaks were detected in the developing endosperm. The first and more prominent peak, at 2 days after flowering, included expression of almost all the tubulin genes. These tubulins are thought to be involved in mitoses during the formation of the syncytial endosperm. The second, less pronounced but more extended, peak included only some of the tubulin genes (HvTUA3, HvTUB1 and HvTUG) and might be associated with the cell wall organization in aleurone and starchy endosperm. The HvTUA5 gene is expressed only in embryo of the developing grain and may be associated with shoot establishment. The expression profiles of the tubulin folding cofactors HvTFC A and HvTFC B as well as small G-protein HvArl2 genes were almost perfectly correlated with the global levels of tubulin mRNA, implying that they have a role in the control of the polymerization of alpha/beta-tubulin heterodimers.