Browsing by Author "Stange, C"
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- ItemTobamovirus coat protein CPCg induces an HR-like response in sensitive tobacco plants(2005) Ehrenfeld, N; Cañón, P; Stange, C; Medina, C; Arce-Johnson, PWhen inoculated into sensitive tobacco Xanthi-nn plants, the crucifer and garlic-infecting Tobacco mosaic virus (TMV-Cg) induces local necrotic lesions that resemble those seen in the hypersensitive response (HR) of resistant tobacco plants. However, unlike these, tobacco Xanthi-nn plants do not become resistant to infection and the virus spreads systemically causing a severe disease characterized by necrotic lesions throughout the plant. To identify the viral protein that elicits this necrotic response, we used a set of hybrid viruses constructed by combination of TMV-Cg and the tobacco mosaic virus strain U1 (TMV-U1). In this study we present evidence that the coat protein of TMV-Cg (CPCg),is the elicitor of the necrotic response in tobacco Xanthi-nn plants. Local and systemic necrotic lesions induced by TMV-Cg and by the hybrid U1-CPCg -that carries CPCg in a TMV-U1 context- are characterized by cell death and by the presence of autoflorescent phenolic compounds and H2O2, just like the HR lesions. In addition, defense-related genes and detoxifying genes are induced in tobacco Xanthi-nn plants after TMV-Cg and U1-CPCg inoculation. We postulate that in our system, CPCg is recognized by sensitive tobacco plants that mount an incomplete defense response. We call this an HR-like since it is not enough to induce plant resistance.
- ItemIdentification and characterization of a novel tobacco mosaic virus resistance N gene homologue in Nicotiana tabacum plants(2004) Stange, C; Matus, JT; Elorza, A; Arce-Johnson, PNicotiana tabacum cv. Xanthi nn plants are susceptible to infection by most tobamoviruses (TMV). However, such plants display a partial hypersensitive resistance response (HR-like response) to TMV-Cg. The genetic mechanism of the HR-like response has yet not been determined, but it may involve a gene with a function similar to that of a resistance gene, responsible for HR in resistant plants. We have cloned a gene homologous to the resistance N gene, named NH, from Nicotiana Xanthi nn plants. The coding region of NH is 5.028 base pairs ( bp) long and has 82.6% nucleotide identity with the N gene. In contrast to the N gene, the NH gene lacks intron 4 and does not have sites for alternative splicing of intron 3. Analysis of its sequence revealed that NH belongs to the TIR/NSB/LRR gene class. We were able to detect stable levels of NH-transcript in Nicotiana Xanthi nn plants from 0 to 18 h post-inoculation (hpi) with TMV-Cg. Transcript levels increased slightly at 24 hpi and dropped below basal values at 48 hpi. The NH transcript was also detected in a range of resistant Nicotiana plants ( N. tabacum Xanthi NN,N. glutinosa, N. glauca and N. rustica) suggesting that NH is a homologue of the N gene, rather than an allele. We have cloned and characterised the NH gene ( GenBank acc. no. bankit598573 AY535010) from nn susceptible plants and postulate that this gene might be involved in the HR-like response seen in these plants.
- ItemIsolation of Pinus radiata genomic DNA suitable for RAPD analysis(1998) Stange, C; Prehn, D; Arce-Johnson, P
- ItemOptimization of in vitro culture conditions for Pinus radiata embryos and histological characterization of regenerated shoots(1999) Stange, C; Prehn, D; Gebauer, M; Arce-Johnson, PDifferent in vitro culture conditions were tested on Pinus radiata organogenic embryos. Optimum shoot induction occurred at 26.1 degrees C, whereas the best elongation resulted at 21.4 degrees C. Supplements of 2.5 mg/l or 5 mg/l of BAP added to the induction media produced a similar number of regenerated shoots, which differed statistically from 1.0 mg/l of BAP and 0.025 mg/l TDZ. Addition of 10 mg/l MnSO4 to LP1/2 medium significantly increased the number and quality of in vitro regenerated shoots. The removal the apical region of shoots cultured in LP 2.5 mg/l of BAP increased the number of de novo generated shoots by 23%, compared to a control group with intact shoots. Approximately 70% of the in vitro shoots of P. radiata were of wet phenotype (hyperhydrated appearance); the rest were waxy in appearance. Histological cuts did not produce any differences in phenotypes, but scanning electronic microscopy of needles gave evidence of differences in epicuticular wax deposits. Abbreviations: LP: Quoirin and LePoivre basal medium, without plant growth regulators; LP,: LP medium + 1 mg/l BAP; LP2.5: LP medium+ 2.5 mg/l BAP; LP5 : LP medium + 5 mg/l BAP; LP1/2: LP basal medium at half strength of macroelements, 2%; commercial sugar, ammonium nitrate 100 mg/l, calcium nitrate 564.5 mg/l, hydroxyquinoleine 1.25 mg/l, MS vitamins and without plant growth regulators; LPT0.025: LP medium + 0.025 mg/l TDZ; BAP: N-6 benzylaminopurine; TDZ: Thidiazuron.
- ItemPhosphorylation of nuclear proteins directs binding to salicylic acid-responsive elements(1997) Stange, C; Ramirez, I; Gomez, I; Jordana, X; Holuigue, LThe cis-located DNA sequence as-1 (Activation Sequence-1) from CaMV 35S promoter has been previously identified as an element that can confer inducibility by salicylic acid (SA) with immediate early kinetics. This sequence specifically binds to ASF-1 (Activation Sequence Factor-1), previously characterized in tobacco nuclear extracts. To assess whether modulation of ASF-1 binding activity can explain the activation of the as-1 sequence observed in vivo, we performed electrophoretic mobility shift assays using nuclear extracts from SA-treated and water-treated tobacco plants. Our results indicate that treatment of plants with SA increases ASF-1 binding to as-1 and to ocs, an as-1-like element from the Agrobacterium octopine synthase gene. In contrast, SA treatment has no effect on the binding of GT-1 factor to its target light-inducible box II element. Furthermore, treatment of nuclear extracts from SA-treated plants with alkaline phosphatase decreases ASF-1 binding to the as-1 element. This can be reversed by pretreatment with 10 mM NaF. Accordingly, pretreatment of nuclear extracts from control water-treated plants with ATP produces an increase in ASF-1 binding activity similar to that observed with SA. This effect of ATP is reversed by treatment with alkaline phosphatase and prevented by quercetin, a casein kinase II inhibitor. These results support the hypothesis that a nuclear protein kinase is involved in the immediate early events of transcriptional activation triggered by SA.
- ItemRegeneration of whole plants from apical meristems of Pinus radiata(2003) Prehn, D; Serrano, C; Mercado, A; Stange, C; Barrales, L; Arce-Johnson, PA methodology to regenerate whole plants of Pinus radiata from apical meristems of 3- and 7-year-old trees was developed. Meristematic domes with two or three leaf primordia were excised from surface-sterilized branch tips of field-grown plants and cultured in LP medium with half strength macronutrients (1/2 LP) and full strength micronutrients. The early growth of meristems required approximately 12 weeks, including a recovery stage during the first 2 weeks. After 8 weeks, some meristems developed abnormal phenotypes and died during the subsequent stages of development. However, healthy meristems elongated and formed shoots when they were transferred to LP medium supplemented with MS vitamins, 30 mg l(-1) casein hydrolysate, and 0.4 g l(-1) agar plus 2.85 g l(-1) Gelrite. Meristems that developed vigorous shoots were used for rooting experiments when they were 2 cm in length. Whole plants were obtained after 5 days of root induction in water-agar medium containing 8.2 muM IBA and 5.4 muM NAA and 1 month culture in LP medium with 10 g l(-1) sucrose. Plants regenerated from meristems were further propagated by rooting of cuttings. Of the rooted cuttings, 10% were morphologically juvenile.