A simple and reliable protocol for regeneration of okra through somatic embryogenesis from suspension cultures has been developed. Embryogenic callus was obtained from hypocotyl explants cultured on media with Murashige and Skoog (MS) salts, Gamborg (B5) vitamins, 2.0 mg dm^-3 2,4-dichlorophenoxyacetic acid (2,4-D), 1.0 mg dm^-3 naphthaleneacetic acid (NAA), 25 mg dm^-3 polyvinylpyrrolidone and 30 g dm^-3 sucrose. More number and high frequency of healthy embryoids appeared individually in suspension culture containing MS salts, B5 vitamins, 2.0 mg dm^-3 2,4-D and 1.0 mg dm^-3 kinetin. Formation of cell clusters from the single cells was clearly noticed during ontogeny. Matured embryos at the cotyledonary stage were transferred to agar solidified medium for germination. The best conversion of embrya into plantlets (67.3 %) was recorded on media with half strength MS salts, B5 vitamins, 0.2 mg dm^-3 benzylaminopurine (BAP) and 0.2 mg dm^-3 gibberellic acid (GA₃). The plantlets were transferred to soil and hardened in the plastic pots. After proper acclimatization, the plantlets regenerated through somatic embryogenesis were compared to seed grown plants to observe any variation.

The effect of elevated carbon dioxide concentration on the changes in the biomass, photosynthesis and nutrient composition was investigated in two leafy vegetables. Spinach (Spinacia oleracea L.) and fenugreek (Trigonella foenum-graecum L.) plants were grown in open top chambers under either ambient (ACO₂, 350 ± 50 µmol mol^-1) or elevated (ECO₂, 600 ± 50 µmol mol^-1) CO₂ concentration and analyzed 40, 60 and 80 days after exposure. The plants grown in ECO₂ had higher net photosynthetic rate and lower stomatal conductance when compared with the plants grown in ACO₂. ECO₂ also changed the nutrient composition: a lower N, Mg and Fe contents and higher C and Ca contents were observed in the leaves of plants exposed to ECO₂ than in those grown at ACO₂.

Shoot apices of Pinus roxburghii Sarg were cultured on Murashige and Skoog's medium (MS) supplemented with cytokinins [6-benzyladenine (BA), kinetin and N-benzyl-9-(2-tetrahydropyranyl) adenine (BPA)] alone and in combination with auxin, α-napthaleneacetic acid (NAA). Of the three cytokinins tested at varying concentrations, medium supplemented with 10 µM BA was found optimal in respect of explant responsiveness (97.22 %) and average number of buds induced per explant (7.42). The concentration of cytokinins in the induction medium had a profound effect on rate of elongation of induced buds on MS basal medium containing 0.5 % activated charcoal. Further, shoots induced on lower concentrations of BA increased up to 2.4 times in length in 4 weeks. Decapitation of the explant enhanced the rate of axillary bud elongation. Proliferating shoot cultures were established by sub-culturing the axillary shoots on MS supplemented with 10 µM BA. Shoots 2-3 cm in length were suitable for culturing as more buds were induced on them compared to longer or shorter shoots. Root primordia were induced on 70.83 % shoots when transferred to 1/2 MS medium supplemented with 5.0 µM NAA. Elongation of root primordia (60 %) was achieved in liquid 1/2 MS basal medium. The plantlets were successfully transferred to soil after hardening; the time period from initiation of shoot buds to transplantation being 20-22 weeks.

The antioxidant defense system in three ecotypes of reed (Phragmites communis Trin.), swamp reed (SR), dune reed (DR), and heavy salt meadow reed (HSMR), from northwest China were investigated. The HSMR possessed the highest ratio of ascorbate (ASC)/dehydroascorbate (DHA) and activities of superoxide dismutase (SOD) and catalase among the three reed ecotypes, whereas, the DR exhibited the highest ratio of glutathione/glutathione disulfide and activities of ASC peroxidase (APX) and DHA reductase. Malondialdehyde and hydrogen peroxide contents were highest in HSMR, intermediate in SR, and lowest in DR. In addition, different isoenzymes of glutathion reductase, APX, SOD and DHA were also observed in three reed ecotypes.

Wheat (Triticum aestivum L. Sakha 61) grains were exposed to microwave radiation of a wavelength 2.85 cm and frequency 10.525 GHz for 15, 45, or 75 min. The exposed grains were germinated and then harvested after 7 and 14 d. While 15-and 45-min exposure of the grains stimulated seedling shoot length and fresh and dry masses, the exposure for 75 min had no pronounced effects. 15 and 75-min irradiation increased succulence and pigment contents in 7-and 14-d-old seedlings. While the ratios chlorophyll (Chl) a/b, carotenoids (Car)/Chl were higher in 7-d-old seedlings than in the control ones for all doses, they decreased in 14-d-old seedlings. The microwave radiation increased protein and amino acid contents, but decreased the contents of saccharides, nucleic acids, and phenolic compounds. Low dose (15 min) stimulated proline synthesis, whereas the other doses showed negative effect on its production.

Epicotyl segments of kumquat (Fortunella crassifolia Swingle cv. Jindan) were transformed with Agrobacterium tumefaciens GV3101 harboring neomycin phosphotransferase gene (npt II) containing plant expression vectors. Firstly, the explants were cultured in darkness at 25 °C on kanamycin free shoot regeneration medium (SRM) for 3 d, and then on SRM supplemented with 25 mg dm^-3 kanamycin and 300 mg dm^-3 cefotaxime for 20 d. Finally, they were subcultured to fresh SRM containing 50 mg dm^-3 kanamycin monthly and grown under 16-h photoperiod. Sixty five kanamycin resistant shoots were regenerated from 500 epicotyl explants after four-month selection. Shoot tips of 20 strong shoots were grafted to 50-day-old kumquat seedlings and survival rate was 55 %. Among the 11 whole plants, 3 were transgenic as confirmed by Southern blotting. This is the first report on transgenic kumquat plants, and a transformation efficiency of 3.6 % was achieved.

Dehydrins present a distinct biochemical group of late embryogenesis abundant (LEA) proteins characterised by the presence of a lysine-rich amino acid motif, the K-segment. They are highly hydrophilic, soluble upon boiling, and rich in glycine and polar amino acids. It is proposed that they can act as emulsifiers or chaperones in the cells, i.e., they protect proteins and membranes against unfavourable structural changes caused by dehydration. Cold usually precedes freezing in nature and induces many physiological and biochemical changes in the cells of freezing-tolerant plant species (cold-acclimation) that enable them to survive unfavourable conditions. It is demonstrated that the induction of dehydrin expression and their accumulation is an important part of this process in many dicotyledons (both herbaceous and woody species), and also in winter cultivars of cereals, especially wheat and barley. Some mechanisms which are proposed to be involved in regulation of dehydrin expression are discussed, i.e., endogenous content of abscisic acid, homologues of Arabidopsis C-repeat binding factor (CBF) transcriptional activators, the activity of vernalization genes and photoperiodic signals. Finally, we outline some new approaches emerging for the solution of the complex mechanisms involved in plant cold-acclimation, especially the methods of functional genomics that enable to observe simultaneously changes in the activity of many genes and proteins in a single sample.

The effect of short-term exposure to elevated CO₂ concentration and high irradiance on the activity of superoxide dismutase (SOD), ascorbate peroxidase (APX), guaiacol peroxidases (GPX) and catalase (CAT), and on the extent of the lipid peroxidation was studied in bean (Phaseolus vulgaris L.) plants. Plants were exposed for 4 d (8 h a day) to irradiance of 100 (LI) or 1000 (HI) μmol m^-2 s^-1 at ambient (CA, 350 μmol mol^-1) or elevated (CE, 1300 μmol mol^-1) CO₂ concentration. Four-day exposure to CE increased the leaf dry mass in HI plants and RuBPC activity and chlorophyll content in LI plants. Total soluble protein content, leaf dry matter and RuBPC activity were higher in HI than in LI plants, although the HI and CE increased the contents of malonyldialdehyde and H₂O₂. Under CA, exposure to HI increased the activity of APX and decreased the total SOD activity. Under CE, HI treatment also activated APX and led to reduction of both, SOD and GPX, enzymes activities. CE considerably reduced the CAT activity at both irradiances, possibly due to suppressed rate of photorespiration under CE conditions.

Inter simple sequence repeat (ISSR) marker assay was employed to validate the genetic fidelity of Swertia chirayita plantlets multiplied in vitro by axillary multiplication upto forty-two passages. Sixteen ISSR primers generated a total of 102 amplicons among the tissue-cultured plants. Forty-eight amplicons were amplified in the outlier (a Swertia species). The outlier (negative control) was employed to rule out the possibility that the invariant fingerprint was due to chance alone and that the ISSR technique employed was not discriminatory enough to detect the off-types. A homogenous amplification profile was observed for all the micropropagated plants. The results confirmed the clonal fidelity of the tissue culture-raised S. chirayita plantlets and corroborated the fact that axillary multiplication is the safest mode for multiplication of true to type plants.

The heat stress-induced dehydrin proteins (DHNs) expression and their relationship with the water relations of sugarcane (Saccharum officinarum L.) leaves were studied. Sugarcane seedlings were subjected to heat stress (day/night temperature of 40/35 °C) under relative humidity 60/65 % to avoid aerial desiccation and determinations made at 4, 12, 24, 36, 48, 60 and 72 h. The leaves showed a sharp decline in the water and osmotic potentials, and relative water content during first 12 h of heat stress, but a regain in their values in 24 h. The pressure potential (ψ_p) decreased initially but increased later and approached control leaves. The increase in ψ_p was tightly correlated to the accumulation of free proline, glycinebetaine and soluble sugars, indicating their possible involvement in the osmotic adjustment under heat stress. Immunological detection revealed the expression of three DHNs with an apparent molecular mass of 21, 23 and 27 kDa under heat stress (48 to 72 h) and their expression was independent of the changes in the water relations of leaves.

The relationship between photosynthetic rate and yield formation processes of the newer and older maize hybrids were investigated. Leaf area at flowering (source) and kernel number (sink) of the newer hybrids were greater than the older ones although their light-saturated photosynthetic rate (P_sat) were not greater than the older ones before flowering. After flowering, P_sat and chlorophyll content of the newer hybrids declined more slowly than the older ones. They not only distributed almost all photosynthates produced after flowering to grain but also reallocated some reserved photosynthates produced before flowering to grain. The newer hybrids exhibited greater grain mass than the older ones mostly because they could optimally regulate the photosynthetic rate and yield formation processes to maximize grain mass.

Cytophysiological and cytogenetic characteristics of cell suspension culture of Panax japonicus var. repens were studied in relation to the accumulation of ginsenosides (GSs). The minimal time of cell number doubling was 1.3 ± 0.1 d and cell number increased 7 to 8-fold during growth cycle. The cell culture can be considered as aneuploid with about tetraploid (46-60 chromosomes) modal class. Upon long-term cultivation, the total content of GSs considerably increased and maximal concentration of GSs was 2.2 %(d.m.). The ratio of seven major GSs only slightly altered both over each and different subcultures. The overall amount of GSs of Rg-group significantly exceeded that of Rb-group. Cell volume and the number of large cellular aggregates with the higher proportion (by 20 %) of parenchymal cells increased late in the subculture. In this time the population contained about 20 % of the cells with doubled amount of nuclear DNA and accompanied with elevation in the GS content. These data prompted us to suggest that biosynthesis of GSs has a link with cell differentiation.

Differential mRNA display was used to identify pathogen-responsive, stress-related genes in potato cell suspensions treated with salicylic acid and a cell wall-derived elicitor from Phytophthora infestans. Among the positive clones identified, one was found to be expressed at a significantly higher level in elicited cells than in control cells. DNA sequencing of this amplicon revealed high homology and identified it as a potato cyclophilin cDNA. The maximum amount of the cyclophilin mRNA was found 9 to 12 h after elicitation. Cyclophilin (CyP) mRNA synthesis was also up-regulated from 12 to 24 h in potato leaves locally infected with zoospores from Phytophthora infestans. However, untreated leaves responding systemically to the pathogen showed only a weak, delayed response at 24 h post infection. The observed accumulation of potato CyP mRNA in response to salicylic acid, P. infestans elicitor and P. infestans infection, suggest that CyPs play an important role in plant stress responses.

Cytokinins N⁶-benzyladenine (BA) and 1-(2-chloropyridin-4-yl)-3-phenylurea (4PU-30) delayed the senescence of detached leaves (3^rd to 7^th leaf node) of wild and ethylene insensitive eti5 mutant of Arabidopsis thaliana. The novel anticytokinins, structural analogues of purine and phenylurea cytokinins also affected the senescence of detached rosette leaves of A. thaliana. They diminished to a significant extent the cytokinin-induced delay of chlorophyll destruction, but without a considerable difference in their action against both types of cytokinins. These results correlated with changes observed in ribonuclease (RNase) activity.

To study the defense mechanism of the resistance to the disease of Chinese wild Vitis species and offer powerful bases for the molecular breeding of highly disease-resistant grape cultivars, using mRNA differential display reverse transcript-PCR (DDRT-PCR) and RACE, a full-length cDNA was isolated from Chinese wild Vitis pseudoreticulata clone Baihe-35-1 inoculated with Uncinula necator by pressing infected leaves under natural field conditions. The cDNA designated as GLOXrg is 1708 bp in length with an open reading frame of 1572 bp encoding 523 amino acids, containing the conserved domain, glyoxaloxid N domain (pfam07250). No homologous nucleotide sequence was found, but the deduced amino acid sequence of GLOXrg shows 74 % identity with a putative protein from Arabidopsis thaliana (accession No. CAB88357). GLOXrg was cloned into pGEX-4T-1 vector. The recombinant vector expressed an about 83-kDa GST-GLOXrg fusion protein as insoluble inclusion bodies in Escherichia coli BL21. Fusion protein GST-GLOXrg was isolated and used to raise the polyclonal anti-(GST-GLOXrg) in rabbits. Western blot analysis showed a high titer (10 000) and the high specificity of the polyclonal anti-(GST-GLOXrg). This is the first report on the cloning of the full-length GLOXrg cDNA from the plant except Arabidopsis thaliana. Through the differential expression, GLOXrg probably plays a crucial role in the resistance to Uncinula necator of Chinese wild Vitis species. An optimized system on prokaryotic expression, protein purification and preparation of the polyclonal antibodies of GLOXrg was also established.

An in vitro method for propagation of Holarrhena antidysenterica Wall. has been developed using nodal explants from mature trees growing in the field. Irrespective of concentrations and combinations of growth regulators used, the axillary and terminal buds sprouted and elongated when inoculated on Murashige and Skoog (MS) medium. The highest numbers of shoots were formed when sprouted shoots were subcultured from MS basal medium onto MS medium containing 2 mg dm^-3 N⁶-benzyladenine (BA) and 0.5 mg dm^-3 α-naphthalene acetic acid (NAA). The shoot number further increased upon subculture on MS medium containing 0.5 mg dm^-3 BA. By repeated sub-culturing of shoots derived from nodal axillary buds, a high frequency multiplication rate was established. The elongated shoots were excised and rooted in auxin free MS basal medium. Ex vitro rooting of in vitro formed shoots was achieved upon dipping the microshoots for 2 min in 2 mg dm^-3 of indole-3-butyric acid solution. Successful field establishment and high (80-90 %) survival of plants was observed.

Two wheat cultivars, HF9703 (drought tolerant) and SN215953 (drought sensitive) were used to examine the effects of glycinebetaine (GB, 100 mM) on lipid composition and function of thylakoid membranes under drought stress. GB application mitigated negative effect of drought on Ca²⁺-ATPase and Hill reaction activities, chlorophyll content, gas exchange and photosynthesis. These positive effects of GB application maybe, in part, correlated with improving the lipid composition of the thylakoid membranes.

Maize (Zea mays L.) seedlings of two cultivars (cv. Bastion adapted to W. Europe, and cv. Batan 8686 adapted to the highlands of Mexico), raised in a glasshouse (19-25 °C), were transferred to 4.5 or 9 °C at photon flux density (PPFD) of 950 µmol m^-2 s^-1 with 10-h photoperiod for 58 h and then allowed to recover at 22 °C for 16 h (14 h dark and 2 h at PPFD of 180 µmol m^-2 s^-1). The ultrastructural responses after 4 h or 26 h at 4.5 °C were the disappearance of starch grains in the bundle sheath chloroplasts and the contraction of intrathylakoid spaces in stromal thylakoids of the mesophyll chloroplasts. At this time, bundle sheath chloroplasts of cv. Batan 8686 formed peripheral reticulum. Prolonged stress at 4.5 °C (50 h) caused plastid swelling and the dilation of intrathylakoid spaces, mainly in mesophyll chloroplasts. Bundle sheath chloroplasts of cv. Batan 8686 seedlings appeared well preserved in shape and structure. Batan 8686 had also higher net photosynthetic rates during chilling and recovery than Bastion. Extended leaf photobleaching developed during the recovery period after chilling at 4.5 °C. This was associated with collapsed chloroplast envelopes, disintegrated chloroplasts and very poor staining.

The organogenic competence of leaf explants of eleven Carthamus species including C. tinctorius on Murashige and Skoog (MS) medium supplemented with different concentrations of thidiazuron (TDZ) + α-naphthaleneacetic acid (NAA) and 6-benzyladenine (BA) + NAA was investigated. Highly prolific adventitious shoot regeneration was observed in C. tinctorius and C. arborescens on both growth regulator combinations and the shoot regeneration frequency was higher on medium supplemented with TDZ + NAA. Nodal culture of nine Carthamus species on media supplemented with BA and kinetin (KIN) individually revealed the superiority of media supplemented with BA over that of KIN in facilitating a higher shoot proliferation index. Proliferating shoots from axillary buds and leaf explants were transferred to medium supplemented with 1.0 mg dm^-3 KIN or 0.5 mg dm^-3 BA for shoot elongation. Elongated shoots were rooted on half-strength MS medium supplemented with 1.0 mg dm^-3 each of indole-butyric acid (IBA) and phloroglucinol. The plantlets thus obtained were hardened and transferred to soil.

The relation between oxygen evolution rate (OER) and quantum yield of photochemical reactions in photosystem 2 (Φ_PS2) was examined in lichen symbiotic alga Trebouxia erici Ahmadjian (strain UTEX 911) exposed to different irradiances and osmotic stress (2 M sucrose for 60 h). Linear relationship was found between OER and Φ_PS2 in control cell suspension within irradiance range of 0 - 500 μmol m^-2 s^-1. Under osmotic stress, OER and Φ_PS2 were significantly reduced. Relation between OER and Φ_PS2 was curvilinear due to strong osmotically-induced inhibition of OER at high irradiance. The highest used irradiance (500 μmol m^-2 s^-1) was photoinhibitory for osmotically-stressed T. erici because non-photochemical quenching (NPQ) increased substantially. Energy-dependent quenching represented major part of NPQ increase. Osmotic stress led also to the reduction of capacity of photochemical processes in PS 2 (F_V/F_M) and increase in F₀/F_M. These changes indicated negative effects of osmoticum on structure and function of photosynthetic apparatus.

Aspects of plant growth such as height, branch number, leaf number, leaf area, pod area, 100-seed mass, etc., were correlated with biochemical changes such as contents of chlorophyll (Chl), proteins, DNA, and RNA, and protease activity during development and senescent phases in leaves, flowers, and pods of Cajanus cajan L. cv. UPAS-120 after treatments with kinetin (Kn). A significant increase was noticed in branch number, leaf number, leaf area, and seed mass while other growth processes registered a small increase after Kn application. Effectiveness of 5 µM Kn was also noticed in minimizing the loss of Chls, proteins, and nucleic acids as well as reducing the protease activity during maturity and senescence. Chl a/b ratio maintained a high value up to 30-d followed by a decline in leaves while flowers registered much lower ratio at 20-d-age. Pods were unique in having relatively lower ratio of Chl a/b in comparison to leaves.

Four rice indica genotypes of local importance were transformed with RC7, rice chitinase cDNA clone through Agrobacterium-mediated gene transfer method using mature seed derived calli as explants. The putative hygromycin resistant calli showed varied level of regeneration efficiency ranging from 2.0 to 7.6 %. The stable integration and expression of RC7 was confirmed through polymerase chain reaction (PCR) and Western analysis. Transformation efficiency ranged from 0.9 to 5.2 %. The expression of RC7 (35 kDa chitinase) in different tissues of transgenic plant (root, sheath and leaf) was proved through Western analysis and in terms of increased chitinase activity. The inheritance of transgene was studied through PCR and Western analysis in transgenic plants of Pusa Basmati 1. Bioassays with transgenic plants of local cultivars exhibited enhanced resistance up to 33.3 % to rice sheath blight pathogen Rhizoctonia solani under glasshouse conditions. Enhanced expression or 3-to 4-fold increased activity of chitinase in transgenic plants was correlated with sheath blight resistance.

Potted 2-year-old lemon trees [Citrus limon (L.) Burm. fil, cv. Verna] grafted on sour orange (C. aurantium L.) rootstock were subjected to flooding for 3 d. Control plants were irrigated daily to field capacity. Continuously (sap flow, trunk diameter fluctuations) and discretely (predawn and midday leaf water potential, leaf conductance) measured plant-based water status indicators were compared. The sensitivity of the maximum daily trunk shrinkage signal intensity to flooding and its behaviour during the recovery period demonstrated that this indicator is more feasible than the others for use in automatic irrigation. The responses to flooding of continuously and discretely measured plant-based water status indicators were very similar to those observed in response to drought stress indicating that it necessary to use soil water measurement automatic sensors to detect the cause of the stress. The results underlined the robustness of the compensation heat-pulse technique for estimating instantaneous and daily transpiration rates on flooding stress and recovery.

Dedifferentiation was monitored in Rubia tinctorum L. leaves over a 14-d period after callus induction using transmission electron microscope (TEM), high performance liquid chromatography (HPLC), spectroscopy and thin layer chromatography (TLC). Photosynthetic pigment loss of leaves took 3-5 d coinciding with the first period of anthraquinone accumulation. Callus cells were discernible in the region of the vascular bundles and wounded edges of leaves after 10-14 d. Characteristic ultrastructural alterations were manifested in vacuolization, appearance of mitochondria, amount of smooth endoplasmatic reticulum and cytoplasm, caryolympha density of nuclei and cytoplasm content of cells. There were special events in the transfer cells: unequal divisions of dedifferentiated plastids and lytic activity in the cell wall. Our results show that mesophyll cells seem to be stopped at a particular level of dedifferentiation, while transfer cells embodied in veins of leaves pass through further alterations and lead to callus formation. Findings suggest that a sort of dedifferentiation drift manifests in the various cells of R. tinctorum leaves during callus induction and depending on their specialized status they achieve different levels of dedifferentiation. Approximately 4 weeks after callus induction, root growth has started from the young calli.

Accumulation of extracellular chitinases in Brassica napus plants infected with Turnip yellow mosaic virus (TYMV) and fungal pathogen Leptosphaeria maculans was studied in both compatible and incompatible interaction. Analysis of apoplast fluid by means of non-denaturing anodic and cathodic PAGE followed by in-gel detection of chitinase activity revealed a number of chitinase isozymes. TYMV induced 8 acidic and 4 basic isozymes in a systemic way. Except for one acidic and one basic isozyme, all other chitinases were also constitutively present in low amounts in mock inoculated control. In TYMV systemically infected plants, chitinases were detected in leaves expressing symptoms as well as in symptomless ones. Both virulent and avirulent L. maculans isolates induced production of chitinase isozymes in cotyledons in a time dependent manner. Some of them were present in plants constitutively and their content increased after inoculation. Three of five acidic and two of three basic isozymes responded to L. maculans infection. Chitinases started to accumulate before symptom appearance. First two acidic isozymes were detected 24 h after inoculation. The difference between compatible and incompatibe interaction reflected two basic isozymes.

Foliar sprays of water or 1, 10 and 100 µM aqueous solutions of gibberellic acid (GA₃) or kinetin (KIN) were applied to 40-d-old plants of Nigella sativa (L.) to study their effects on net photosynthetic rate, nitrogen metabolism, and the seed yield. 10 µM solutions of both the hormones, especially GA₃, appreciably increased the activities of nitrate reductase and carbonic anhydrase, chlorophyll and total protein contents and net photosynthetic rate in the leaves, along with capsule number and seed yield plant^-1, at harvest.