Acidic and neutral ribonucleases (RNases) were studied in embryos of Triticum durum cv. Cappelli and the effects of abscisic acid (ABA) and gibberellic acid (GA₃) were analysed. RNases activities increased during germination and were comparable in dormant and non-dormant embryos imbibed for 24 h. ABA generally inhibited ribonucleolytic activities, while GA₃ only affected dormant embryos. To assess whether changes in RNase activities during germination or following hormonal treatment required new transcriptional or translational action, cycloheximide or cordycepin were used. The action of inhibitors of acidic RNase activity was found only in non-dormant-embryos. Findings obtained in the present work concur with a change of the ribonucleolytic pattern in the shift from dormant to non dormant metabolism.

Wheat (Triticum aestivum L.) cvs. HD 2285 (relatively tolerant) and WH 542 (susceptible) were exposed to ambient and elevated temperature (3-4 °C higher) in open top chambers during post anthesis period. The grain yield components were determined at the time of maturity. In order to elucidate the basis of differential tolerance of these cultivars, the excised developing grains (20 d after anthesis) of ambient grown plants were incubated at 15, 25, 35 and 45 °C for 2 h and then analysed for the activities of soluble starch synthase (SSS), granule bound starch synthase (GBSS), kinetic parameters of SSS and content of heat shock protein (HSP 100). The elevated temperature during grain development significantly decreased grain growth in WH 542 whereas no such decrease was observed in HD 2285. High temperature tolerance of HD 2285 was found to be associated with higher catalytic efficiency (V_max/K_m) of SSS at elevated temperature and higher content of HSP 100.

The Agrobacterium-mediated transformation was done in rice (Oryza sativa L. var. indica) cv. HKR126 and elite cross-bred cv. Pusa Basmati1 (PB1), using strain LBA4404 containing pCAMBIA1300 cloned with gene cassettes; potato proteinase inhibitor and Bacillus thuringiensis endotoxin (plasmid JDW53) or mannitol-1-phosphate dehydrogenase (plasmid RKJ108). Co-cultivation with scutellar-calli derived from mature seeds showed stable and highly efficient transformation. In cvs. HKR126 and PB1, 35 % and 41 % of hygromycin resistant calli were obtained. The transformation efficiency in PB1 (22.0 %) was much higher than in HKR126 (12.5 %). Similarly, PB1 had higher plant regeneration efficiency than HKR126. The shoots regenerated per callus were, 3-4 in HKR126 and 5-6 in PB1. The transformation efficiency with pRKJ108 (18.6 %) was higher than pJDW53 (15.9 %). Polymerase chain reaction (PCR) analysis showed the presence of transgenes in regenerated transgenic plants of both cultivars.

Systemic acquired resistance (SAR) can be induced in plants by incompatible pathogens, pathogen derived extracts, or certain chemicals as benzothiadiazole (BTH). The aim of this work was to compare changes in peroxidase and chitinase activities, enzymes considered as PR-proteins, caused by BTH and the pathogen Plasmopara halstedii. Hypocotyls from susceptible and resistant BTH-treated sunflower seedlings showed increased peroxidase and chitinase activities. Inoculation with P. halstedii increased chitinase and peroxidase activities in inoculated hypocotyls from susceptible but not from resistant sunflower seedlings.

Transformation and high efficient regeneration of transgenic plants from embryogenic calluses of Bingtang sweet orange [Citrus sinensis (L.) Osbeck] was reported. Embryogenic calluses were inoculated with Agrobacterium tumefaciens strain EHA105, harboring the binary Ti plasmid pROK II and carrying a neomycin phosphotransferase II (NPTII) gene, an intron β-glucuronidase (GUS) gene and the Arabidopsis APETALA1 (AP1) gene. Transformation treatment was with inoculation time of 30 min, co-culture of 3 d at 23 °C and supplementation of the co-culture medium with 2 mg dm^-3 acetosyringone (AS). Kanamycin (50 mg dm^-3) was effective to inhibit the growth of non-transformed calluses while it did not affect the transformed ones. The total number of transformed callus lines was 7 with 100 % embryo induction. High efficient regeneration of the transgenic embryos (88 % with 4-5 shoots per embryoid) was realized within 3 months. Integration of the transgene into the citrus genome was confirmed by histochemical GUS staining, polymerase chain reaction (PCR) analysis with AP1-specific primer and Southern blot hybridization with a 712 bp PCR fragment of AP1 as the probe.

Allium cepa L. roots were exposed to 0.1 and 1.0 mM Cd for 6, 24 and 48 h and the localization of Cd in the root tissue was investigated. Scanning electron microscopy (SEM) and energy dispersive X-ray analysis (EDXA) were performed on frozen-dried tissues of roots. No Cd was detected in the roots treated with only 0.1 mM Cd, while after exposure to higher Cd concentration (1.0 mM) Cd was observed in cell wall and in cytoplasm in the epidermis, cortex and vascular tissues in the roots.

Water relations, mineral composition, growth and root morphology were studied in pepper plants (Capsicum annuum L. cv California Wonder). Two NaCl concentrations (30 and 60 mM) and two nutrient solutions in which the concentrations of macronutrients were increased were used to assess the ionic and osmotic effects of NaCl in these plants. The hydraulic conductivity (L_o), stomatal conductance (g_s), percentage of open stomata and pressure potential (Ψ_p) decreased with all treatments, in a similar way for 30 mM NaCl and for its iso-osmotic solution of macronutrients, however, the decrease was higher for 60 mM NaCl than for its iso-osmotic solution. Ion analyses also revealed that nutrient concentrations were altered greatly at 60 mM NaCl. Also, changes in morphology, such as increases in cortex cell size and in intercellular spaces, were detected. Therefore, at low salinity, the effect of NaCl was mainly osmotic, however, under higher salinity also the toxicity of Na⁺ and Cl^- participate.

Changes in growth and structural properties of Citrus cell line Carvalhal acclimated to 100 mM NaCl in the medium were compared to unacclimated control cells and cells exposed to 100 mM NaCl. Transmission electron microscopy (TEM) showed presence of ring-shaped mitochondria, increase in the number of amyloplasts and lipid bodies, higher cell wall thickness and partitioned vacuoles in acclimated cells.

A method for plant regeneration in Robinia pseudoacacia L. from cell suspension culture was established. Non regenerative friable callus from hypocotyls and cotyledon explants from in vitro raised seedling induced on solid Murashige and Skoog (MS) medium supplemented with 0.05 mg dm^-3 2,4-dichlorophenoxyacetic acid (2,4-D) was used for initiation of cell suspension cultures on same MS medium but without agar. Single cells were isolated after 3 d and the optimum cell density was 1-3 × 10⁴ cells per cm³ of the liquid MS medium. Plating efficiency was 29.6 % and callus formed within 4 weeks was subcultured and transferred to solid MS medium supplemented with 0.6 mg dm^-3 benzyladenine (BA) along with 0.05 mg dm^-3 α-naphthalene-1-acetic acid (NAA) for the induction of adventitious bud primordia. The shoots developed were isolated and re-cultured on MS medium containing 0.6 mg dm^-3 BA. These microshoots after dipping in 1-2 cm³ of 10 mg dm^-3 indole-3-butyric acid (IBA) for 24 h in dark were cultured on half strength solid MS medium supplemented with 0.05 % charcoal and showed 80-82 % rooting within 4 weeks.

Pericarp polypeptide profiles were analyzed at three ripening stages in the F₁ hybrid and the F₂ population from the cross between the accessions: LA1385 (Lycopersicon esculentum var. cerasiforme) and 804627 (L. esculentum, a homozygous genotype for the nor mutant). Six polymorphic polypeptides were observed in LA1385, while no polymorphic polypeptides among ripening stages was observed in 804627. On the other hand, some polypeptides in the F₁ hybrid were not observed in the parents whereas others were present in both parental genotypes and were unnoticeable in the hybrid genotype. From a cluster analysis on the protein profiles of the F₂ population, the differential expression of proteins allowed to distinguish mature green (MG) stage from the others two stages, while for breaker stage (BR) and red ripe stage, the genetic background was more important in forming groups. The differential expression of proteins could be associated with fruit morphology traits such as a 72 kDa polypeptide present in MG stage with fruit diameter, height and mass and a 47 kDa polypeptide found in BR with fruit shelf life.

A set of 30 accessions of five Curcuma species-C. latifolia, C. malabarica, C. manga and C. raktakanta and 13 morphotypes (identified on the basis of morphological markers) of C. longa conserved in the In Vitro Genebank at National Bureau of Plant Genetic Resources, New Delhi, were subjected to RAPD analysis. Of the 200 RAPD primers screened, 21 polymorphic primers were selected for further study. Mean genetic similarities based on Jaccard's similarity coefficient ranged from 0.18 to 0.86 in accessions of cultivated species, i.e., C. longa and from 0.25 to 0.86 in wild species. The dendrogram derived from the RAPD data corroborated the morphological classification of the morphotypes. The efficiency of individual RAPD primers was also compared; primers OPC-20, OPO-06, OPC-01 and OPL-03 were adjudged highly informative in discriminating the germplasm of Curcuma.

In this study, Dendrobium Sonia 17 plantlets were used to induce in vitro flowering. Inflorescences were induced and rooting was inhibited in the half-strength Murashige and Skoog medium containing 20 µM N ⁶-benzyladenine (BA). The medium with high P and low N contents was effective to induce inflorescences while the medium with low P and high N contents was only effective to promote forming of shoots. In addition, the induced in vitro inflorescences were able to multiply and maintain without exhibiting a distinctive vegetative phase. Different morphologies of in vitro flowers such as incomplete flower structures, abnormal and unresupinated in vitro flowers were observed.

Proline accumulation was studied in the leaves of Glycine max (L.) Merr. subjected to salt stress in the presence of aminoguanidine (AG, a specific inhibitor of diamine oxidase, DAO) and exogenous putrescine (Put). Both DAO activity and proline content were increased while endogenous Put content was decreased in soybean leaves under 50 to 150 mM NaCl. There was a negative correlation between proline accumulation and endogenous Put content. The addition of AG during NaCl stress inhibited DAO activity, caused Put accumulation and a 15 to 20 % decrease in proline content. Application of 1 mM Put to NaCl solution markedly increased proline content. The promotive effect of Put application could be alleviated by the treatment with Put plus AG. Moreover an application of AG had no effect on proline accumulation in soybean seedlings grown under normal condition. These results indicate that the quantitative contribution of Put degradation to proline formation is 15 to 20 %.

An in vitro propagation system was developed for castor-bean (Ricinus communis L. cv. TMV 6) through cotyledon derived callus cultures. The impact of different concentrations of auxins, cytokinins, additives, amino acids and sugars were evaluated for callus induction and shoot proliferation. Green compact nodular organogenic callus was obtained on the medium fortified with Murashige and Skoog (MS) salts, B5 vitamins, 2.0 mg dm^-3 6-benzyladenine and 0.8 mg dm^-3 α-naphthalene acetic acid (NAA). Multiple shoot proliferation from the callus cultures was achieved on the medium with MS salts, B5 vitamins, 2.5 mg dm^-3 thidiazuron (TDZ), 0.4 mg dm^-3 NAA and 15 mg dm^-3 glutamine. During multiple shoot induction the phenolic secretion was controlled by the addition of 15 mg dm^-3 polyvinylpyrolidone. The proliferated shoots were elongated on the medium comprising MS salts, B5 vitamins, 1.5 mg dm^-3 TDZ and 0.3 mg dm^-3 gibberellic acid. The elongated shoots were rooted on the medium containing MS salts, B5 vitamins, 0.3 mg dm^-3 indole-3-butyric acid and 0.6 mg dm^-3 silver nitrate. After root induction, the plants were hardened in earthen pots containing sand, soil and vermiculite.

The effects of nitrogen fertilization on the growth, photosynthetic pigment contents, gas exchange, and chlorophyll (Chl) fluorescence parameters in two tall fescue cultivars (Festuca arundinacea cv. Barlexas and Crossfire II) were investigated under heat stress at 38/30 °C (day/night) for two weeks. Shoot growth rate of two tall fescue cultivars declined significantly under heat stress, and N supply can improved the growth rates, especially for the Barlexas. Chl content, leaf net photosynthetic rate, stomatal conductance, water use efficiency, and the maximal efficiency of photosystem 2 photochemistry (F_v/F_m) also decreased less under heat stress by N supply, especially in Crossfire II. Moreover, cultivar variations in photosynthetic performance were associated with their different response to heat stress and nitrogen fertilization, which were evidenced by shoot growth rate and photosynthetic pigment contents.

Pepper (Capsicum annuum L.) plants were sprayed with salicylic acid (SA) and treated with ultraviolet radiation UV-A (320-390 nm), UV-B (312 nm), and UV-C (254 nm) of 6.1, 5.8, and 5.7 W m^-2, respectively. UV significantly reduced contents of chlorophyll (Chl) a and b, and carotenoids (Car). SA treatment moderated Chl and Car reduction in plants treated with UV-B and UV-C. The quantity of antocyanins, flavonoids, rutin, and UV-absorbing compounds in plants that were treated with UV-B, UV-C, and SA were significantly increased. Foliar spray of SA counteracted the UV effects on pepper.

In higher plants, fatty acid hydroperoxides are intermediates in the synthesis of a diverse group of bioactive compounds. We used the reverse-transcriptase polymerase chain reaction to isolate a gene responsible for the oxidization of fatty acids from Petunia hybrida. A P450 cDNA that has not previously been isolated (CYP76A3) contained an open reading frame predicted to encode a polypeptide consisting of 507 amino acid residues. The cyp76A3 cDNA was expressed in Saccharomyces cerevisiae AH22 cells under the control of an alcohol dehydrogenase promoter and terminator. The recombinant yeast microsome containing the CYP76A3 hemoprotein was found to specifically catalyze ω-hydroxylation of myristic acid. A high level of the transcripts of the cyp76A3 gene was found in the leaves and roots of P. hybrida, but not in the stems and flowers.

This paper describes multiple shoot regeneration from leaf and nodal segments of a medicinally important herb Centella asiatica L. on Murashige and Skoog's (MS) medium supplemented with a range of growth regulators. The highest number of multiple shoots was observed on MS augmented with 3.0 mg dm^-3 N⁶-benzylaminopurine (BAP) and 0.05 mg dm^-3 α-naphthaleneacetic acid (NAA). Leaf explant showed maximum percentage of cultures regenerating shoots (81.6 %), with the highest shoot number (8.3 shoots per explant) and the shoot length (2.1 cm) whereas, nodal explant showed less number of shoots with callus formation at the base cut end. Successive shoot cultures were established by repeatedly sub-culturing the original explant on a fresh medium. Rooting of in vitro raised shoots was best induced on half strength MS supplemented with 0.5 mg dm^-3 indole-3-butyric acid (IBA) with highest percentage of shoot regenerating roots (76.8 %) with 3-4 roots per shoot. Plantlets were acclimated in Vermi-compost and eventually established in soil. Contents of chlorophyll, total sugars, reducing sugars and proteins were estimated in leaf tissue from both in vivo and in vitro raised plants. Chlorophyll content was higher in in vivo plants, whereas other three components were higher in in vitro plants.

In this study, bulked segregant analysis (BSA) was used on Larix leptolepis × Larix olgensis hybrids to identify a random amplified polymorphic DNA (RAPD) marker associated with high rooting ability in larch. Two DNA bulks: H (high rooting ability) bulk and L (low rooting ability) bulk were constructed according to the rooting percentages of the stock plants. Among the 328 primers, only S356 could amplify a specific band, named S356₄₄₅, which only existed in the H bulk and was further confirmed following selective genotyping of individual hybrids. Grounded on the border sequences, S356₄₄₅ was converted to a sequence characterized amplified region (SCAR) marker, HRL445, which can be useful in marker-assisted selection (MAS) to screen for larch with high rooting ability. All the results strongly indicated that S356₄₄₅ and HRL445 were closely associated with high rooting ability in larch.

The effects of silicon application before sowing on the drought-induced oxidative stress and antioxidant defense in wheat (Triticum aestivum L.) were investigated. Drought stress was applied by withholding watering till sampling at booting or filling stage. Application of Si increased the water potential of drought-stressed plants at filling stage, whereas it did not at booting stage. The superoxide dismutase (SOD) activity was inhibited and peroxidase (POD) activity was enhanced by drought at booting stage, and no differences were observed due to the Si treatment. At filling stage, however, application of Si increased the SOD activity and decreased the POD activity of drought-stressed plants. The catalase (CAT) activity was slightly increased by drought only in the absence of Si and at booting stage. The activity of glutathione reductase (GR) was not greatly influenced. Application of Si did not change the contents of H₂O₂, total soluble protein and protein carbonyl of drought-stressed plants at booting stage, whereas at filling stage, it decreased the content of H₂O₂ and protein carbonyl and increased the content of total soluble protein. The content of thiobarbituric acid reactive substances (TBARS) and the activities of acid phospholipase (AP) and lipoxygenase (LOX) in drought-stressed plants were also decreased by application of Si at both stages.

Exposure of Lemna minor L. to high temperatures leads to an initial decrease in the ubiquitin (Ub) monomer pool size and the accumulation of high molecular mass Ub-protein conjugates, possibly reflecting an increment in the supply of protein substrates to the Ub/proteasome pathway. Alternative explanations include, for example, changes in the transcription rates of one or more pathway components. To measure the effect of heat shock on the simultaneous rates of transcription of selected genes encoding five Ub pathway components (Ub, E1, E2, β subunit and ATPase subunit of the 26S proteasome), a semi-quantitative RT-PCR method was developed using cDNA normalized against the housekeeping gene encoding the 18S ribosomal RNA. Whilst Ub transcription is abruptly increased, there is a moderate increment in the transcription of E1 and the β subunit, a moderate reduction in the transcription of the ATPase subunit and a marked reduction in the case of E2, indicating a differential transcription pattern of the various components of the Ub/proteasome pathway in L. minor subjected to high temperatures. These observations suggest that the increment in the Ub/proteasome pathway intermediates is due to an augmented supply of substrates derived from the stress-induced damage imposed on the cellular proteins. The initial build up of intermediates occurs not only at the expense of the pre-existing pool of free Ub, but also as a result of the prompt increase in Ub expression.

Embryo-rescue was used to facilitate interspecific hybridization of Cucumis anguria L. and C. zeyheri Sond. Embryos were excised from developing fruits at one week intervals for six weeks after hand pollination. Medium containing coconut water was the most suitable for initial germination, and a medium with ascorbic acid was the best for embryo development and plant recovery. Viable plants were obtained from embryos and these plants showed morphological characteristics different from both parents. The analysis of the leucine aminopeptidase (LAP) locus revealed three hybrid types, H1.1, H1.2 and H2.

The coat protein gene isolated from Papaya ringspot virus, Thai isolate, was used to generate transgenic papayas. A binary vector containing the coat protein gene under the control of a 35S promoter, was constructed and transformed into somatic embryos of papaya cultivar Khak Dum by microprojectile bombardment. Eight transgenic lines were identified from 1980 bombarded calli of papaya somatic embryos under kanamycin selection. Integration of the transferred genes into kanamycin resistant papaya calli was verified by PCR amplification of the coat protein gene, GUS assays and Southern blot hybridization. Although the coat protein gene was detected in all transgenic lines, only line G2 was found to be highly resistant to virus. This resistant line showed high degree of rearrangement of the inserted coat protein expression cassette while the coat protein gene itself had a deletion of 166 bp on the 3' end of its sequence. Although the transcription of the coat protein gene was detected in all transgenic lines by RT-PCR, only two transgenic papayas expressed the intact coat protein. Moreover, in the resistant line G2 the amount of the truncated coat protein mRNA was significantly decreased. These results point to an RNA mediated mechanism of coat protein mediated resistance in papaya, probably based on post-transcriptional gene silencing.

A micropropagation protocol through multiple shoot formation was developed for Thlaspi caerulescens L., one of the most important heavy metals hyperaccumulator plants. In vitro seed-derived young seedlings were used for the initiation of multiple shoots on Murashige and Skoog (MS) medium with combinations of benzylaminopurine (BA; 0.5-1.0 mg dm^-3), naphthaleneacetic acid (NAA; 0-0.2 mg dm^-3), gibberellic acid (GA₃; 0-1.0 mg dm^-3) and riboflavin (0-3.0 mg dm^-3). The maximum number of shoots was developed on medium containing 1.0 mg dm^-3 BA and 0.2 mg dm^-3 NAA. GA₃ (0.5 mg dm^-3) in combination with BA significantly increased shoot length. In view of shoot numbers, shoot length and further rooting rate, the best combination was 1.0 mg dm^-3 BA + 0.5 mg dm^-3 GA₃ + 1.0 mg dm^-3 riboflavin. Well-developed shoots (35-50 mm) were successfully rooted at approximately 95 % on MS medium containing 20 g dm^-3 sucrose, 8 g dm^-3 agar and 1.0 mg dm^-3 indolebutyric acid. Almost all in vitro plantlets survived when transferred to pots.

The effects of salinity on growth, leaf nutrient content, water relations, gas exchange parameters and chlorophyll fluorescence were studied in six-month-old seedlings of citrus (Citrus limonia Osbeck) and rooted cuttings of olive (Olea europaea L. cv. Arbequina). Citrus and olive were grown in a greenhouse and watered with half strength Hoagland's solution plus 0 or 50 mM NaCl for citrus, or plus 0 or 100 mM NaCl for olive. Salinity increased Cl^- and Na⁺ content in leaves and roots in both species and reduced total plant dry mass, net photosynthetic rate and stomatal conductance. Decreased growth and gas exchange was apparently due to a toxic effect of Cl^- and/or Na⁺ and not due to osmotic stress since both species were able to osmotically adjust to maintain pressure potential higher than in non-salinized leaves. Internal CO₂ concentration in the mesophyll was not reduced in either species. Salinity decreased leaf chlorophyll a content only in citrus.

A high efficient four step protocol (callus initiation, regeneration, shoot elongation and rooting) for in vitro propagation of Dracaena sanderiana Sander ex Mast was developed. Callusing was achieved from nodal stem segment explants treated with various concentrations of ethylmethane sulphonate (EMS) on MS medium supplemented with 2,4-dichlorophenoxyacetic acid (2,4-D; 1.5 g m^-3). A significant increase in callus induction percentage and biomass production was noticed from lower EMS treated lines (ET₁ and ET₂) comparatively to control and other (ET₃, ET₄ and ET₅) lines. Calli of ET₁ line showed high regeneration potential on MS medium with N ⁶-benzylaminopurine (BAP; 1.75 g m^-3). Length of microshoots, which was reduced by EMS, restored by addition of gibberellic acid (GA₃; 0.4 g m^-3). A marked increase in rooting with increasing EMS concentration was noticed on MS medium fortified with 3-indolebutyric acid (IBA; 1.5 g m^-3).

A cDNA sequence OsMT3 was initially isolated from the subtractive cDNA library of ammonium-fed rice (Oryza sativa L.) leaves, which was further confirmed by Northern blot to be highly ammonium-up-regulated as compared to nitrate. Its full-length cDNA was cloned by RT-PCR, and in silico analysis reveals that the cDNA includes an open reading frame of 186 bp and encodes a rice metallothionein type 3 peptide. Northern blotting analysis showed that OsMT3 gene predominantly expressed in rice leaves, weakly in stems, and barely in buds and roots. The gene transcripts in leaves were significantly induced by polyethylene glycol (PEG), low temperature, NaCl and Cu²⁺, but not by Pb²⁺. Activities of three anti-oxidative enzymes (superoxide dismutase, catalase and peroxidases) and two non-enzymic antioxidants (reduced ascorbate and reduced glutathione) little differed in ammonium-and nitrate-fed rice leaves, indicating that the induced OsMT3 expression was not mediated by ammonium-elicited oxidative signals.

In the reproductive organs of maize (Zea mays L.), the changes in transcription that occur during meiosis at early stage of water deficit were characterized. We used oligo microarray analysis, which included 57452 transcripts representing more than 30 000 genes, and combined this with reverse Northern blot analysis. After 1 d stress, 1 809 transcripts were differentially expressed (by 2-fold or greater) with 34 % of them being upregulated in the tassels, while in the ears 861 transcripts were differentially expressed with 41 % being upregulated. Of these, 33 transcripts were upregulated in both organs, including those involved in protective functions, reactive oxygen species detoxification, nitrogen metabolism and gibberellin metabolism. In contrast, the transcripts involved in polyamine and sugar metabolism were downregulated.