A method was devised for infecting Anthurium andraeanum Linden ex André, an economically important ornamental monocot, with Agrobacterium tumefaciens. Tumors were obtained on plant stems 7 to 10 weeks after inoculation with oncogenic A. tumefaciens strains C58 and A281 cultured previously in an induction medium containing 200 μm acetosyringone at pH 5.5. A higher percentage of tumors were formed in vitro on etiolated internodes (32%) than on green leaf (2%) or petiole explants (3%) 4 weeks after inoculation with induced C58. All explants treated with nontumorigenic A. radiobacter or with induction medium alone failed to produce tumors. Chromatograms showed an accumulation of nopaline in internode explant tumors induced with C58. DNA amplification and hybridization studies showed that the DNA from these tumors, but not from noninoculated anthurium tissue, contained sequences homologous to the nopaline synthase gene of A. tumefaciens T-DNA. Chemical names used 3,5-dimethoxy 4-hydroxyacetophenone (acetosyringone).
A.R. Kuehnle and N. Sugii
A.R. Kuehnle and N. Sugii
Leaf explants of seven cultivars of Hawaiian anthuriums (Anthurium andraeanum Linden ex André cv. Kaumana, Kozohara, Marian Seefurth, Mauna Kea, Nitta, Ozaki, and Paradise Pink) produced callus most successfully after 2 to 3 months on a modified Pierik medium containing 0.36 μm 2,4-D and 4.4 μm BA. Petiole explants callused best on Pierik modified Pierik, and Finnie and van Staden media. Long-term cultures of callus from Univ. of Hawaii anthurium selections UH965, UH1060, and UH1003 were maintained for 12 to 13 months and were still capable of plantlet regeneration. Adventitious plantlets were recovered from callus plated on a Kunisaki medium containing 2.2 or 22 μm BA. Regeneration appeared to be organogenic rather than embryogenic and varied among the genotypes tested. Chemical names used: N- (phenylmethyl)-1H-purin-6-amine (BA); (2,4-dichlorophenoxy) acetic acid (2,4-D).
A.R. Kuehnle, T. Fujii, F.C. Chen, A. Alvarez, N. Sugii, R. Fukui, S.L. Aragon, and J.M. Jaynes
Two cultivars of Anthurium andraeanum Hort. hybrids, `Paradise Pink' and `Tropic Flame', were transformed by Agrobacterium to contain gene sequences for Shiva-1, a cecropin-based lytic peptide. The antibacterial gene was driven by a 35-35S cauliflower mosaic viral (35-35S CaMV) promoter and the construct included the secretory signal sequence for pathogenesis-related protein 1b (PR1b). Blight tolerance of regenerated plants was tested by inoculation with a virulent strain of Xanthomonas axonopodis (formerly campestris) pv. dieffenbachiae (Xad) that is bioluminescent to allow detection of symptomless infections in Shiva-1 transformants. Primary regenerants for two Shiva-1 transgenic lines of `Paradise Pink' displayed significantly enhanced tolerance to bacterial blight over blight susceptible `Rudolph' and even the blight tolerant `Kalapana'. Two Shiva-1 transgenic lines of `Tropic Flame' showed no improved resistance when compared to the control at the mean percent leaf infection level. One Shiva-1 transgenic line of `Tropic Flame' was unexpectedly more susceptible to blight than the nontransgenic control. Low expression of Shiva-1 observed in this line is hypothesized to be the cause of its increased susceptibility to Xad.