The effects of proline and/or glycine betaine (GB) application on growth, photosynthetic pigments, H₂O₂ content, and activities of antioxidant enzymes in rice (Oryza sativa L. cv. KDML105) under salt stress were investigated. The H₂O₂ content and the activities of superoxide dismutase (SOD), glutathione reductase (GR), and ascorbate peroxidase (APX) but not catalase (CAT) increased under salinity. Under 160 mM NaCl, the CAT activity was maintained on the pre-stress level in the presence of proline, whereas in the presence of GB, the GR activity increased more than without GB application. A co-application of 30 mM proline and 1 mM GB did not reduce the increase in H₂O₂ caused by the NaCl stress more than applying each of the osmoprotectants and no synergistic effect on the antioxidant enzymes was observed. However, the application of both the osmoprotectants was the most effective in alleviating degradation of photosynthetic pigments.

Strawberry (Fragaria × ananassa) contains anthocyanins which are important secondary metabolites and key contributors to the antioxidant capacity and nutritional value of the fruit. Anthocyanin biosynthetic genes have been identified. However, the detailed mechanism responsible for anthocyanin accumulation and regulation of biosynthetic genes during strawberry fruit ripening remain unclear. In the present study, we examined the effect of a Fragaria × ananassa myeloblastosis 1 homolog, FaMYB1, on anthocyanin accumulation in the strawberry fruit receptacle. Expression analysis shows that FaMYB1 transcripts increased in response to irradiance but not to abscisic acid treatments. Down-regulation of FaMYB1 was achieved in planta using Agrobacterium-mediated RNA interference (RNAi). As a result, FaMYB1-RNAi fruits exhibited a significant increase in anthocyanin content. Conversely, overexpression of FaMYB1 resulted in a decrease in anthocyanin content. Overexpression of FaMYB1 also significantly reduced expression of genes encoding anthocyanidin synthase and flavonoid glycosyltransferase, whereas down-regulation of FaMYB1 resulted in a significant decrease in the amount of transcripts of leucoanthocyanidin reductase. These data suggest that FaMYB1 might negatively control anthocyanin biosynthesis in the strawberry fruit at the branching-point of anthocyanin/proanthocyanidin biosynthesis.

The Arabidopsis thaliana (L.) Heynh. atExt1 extensin gene is expressed in a cell and tissue-specific manner, in response to developmental cues, and is inducible by a wide range of biotic and abiotic stresses. Over-expression of this gene has been shown to alter stem morphology and to limit the invasiveness of virulent bacterial pathogens, indicating that this cell wall protein gene plays an important role in plant development and defense. A detailed sequence analysis of 3.2 kb of the atExt1 gene promoter region has identified a large number of putative 5'cis-acting elements. Based on the location of clusters of putative promoter control elements, seven atExt1 5' promoter truncations were constructed, fused upstream of the β-glucuronidase (GUS) reporter gene, and transformed into A. thaliana. Transgenic plants carrying the various promoter constructs were challenged by wounding and pathogen attack and analysed for GUS expression - this analysis revealed a complex pattern of regulation, involving positive and negative control regions. Northern analysis using wounded tissues from transgenic Arabidopsis plants carrying the 3.2 kb-promoter::GUS construct confirmed the transcriptional activation of the transgene.

ATP binding cassette (ABC) proteins constitute one of the largest families of transporters. In Arabidopsis thaliana, over 100 genes encoding ABC transporters have been identified. Here, we described the expression pattern of AtABCC13/MRP11, a member of the multidrug resistance associated protein MRP/ABCC subfamily, previously uncharacterized. The histochemical analysis of transgenic Arabidopsis harboring AtABCC13 promoter-β-glucuronidase gene fusion shows that the AtABCC13 expression was specifically associated with vascular bundles. Moreover, AtABCC13 displayed a complex hormonal regulation. β-glucuronidase (GUS) fluorimetric assays revealed that the gene expression was induced by gibberellic acid and downregulated by naphthalene acetic acid, abscisic acid, and zeatin. Because AtABCC13 is also expressed during seed development and during germination, its expression was assessed upon exposure to various nutrients: nitrate, phosphate, and sucrose stimulated the AtABCC13 expression in seedlings, whereas their lack strongly reduced it.

To gain a deep understanding of the regulatory mechanism of Cu-responsive microRNAs (miRNAs) in non-heading Chinese cabbage (Brassica campestris ssp. chinensis Makino), the transcription of 10 annotated stress-inducible miRNAs and their target genes were investigated in two cultivars Suzhouqing and Wutacai exposed to excess of copper. Results show that these miRNAs were negatively correlated with their target genes under the Cu stress and showed different transcriptions in different tissues and cultivars. The transcriptions of bra-miR1530a and bra-miR1533v were highest in petioles and lowest in roots. Bra-miR1533ah, bra-miR1533m, bra-miR1533t, bra-miR414a, and bra-miR398b had the highest and lowest transcriptions in leaves and roots, respectively. In contrast, the transcription of bra-miR172f was highest in roots and lowest in leaves. Bra-miR1533aj and bra-miR1533d had similar transcriptions in petioles and leaves. The promoter analysis further revealed that seven miRNAs contained the Cu-response element (CuRE). In addition, miRNAs with more CuREs in the 5'-flanking sequences showed a lower expression following the Cu treatment. It imply that CuREs likely played a role in increasing the response to Cu in non-heading Chinese cabbage.

Litchi (Litchi chinensis Sonn.) is an important tropical and subtropical evergreen woody fruit tree, and it has been shown that nitric oxide (NO) could promote litchi flowering. NO responsive genes of litchi (cv. Nuomici) primordia were identified through a suppression subtractive hybridization (SSH) library screen. We obtained 1 563 expressed sequences tags (ESTs) that were enriched in the NO treated inflorescence primordia. We then used a reverse Northern analysis to identify 728 true NO responsive ESTs, the sequences of which have been further analyzed. They represent 70 litchi unique genes that could be classified into 9 categories: 14 % of them were involved in transport facilitation, 7 % in transcription regulation, 9 % in stress response, 7 % in sugar metabolism, 9 % in secondary metabolism, 10 % in intracellular signalling, and 44 % in other metabolism, whereas 11 % were genes with unknown functions, and 7 % were genes with no hit found. Next, we performed a real-time quantitative polymerase chain reaction (RT-qPCR) to determine the expression of selected candidate genes during a time-course of NO treatment and of normal floral tissue development.

The effects of acclimation to cold (4 °C) and heat (36/38/40 °C) on corresponding freezing and heat tolerances of one-year-old Trigonobalanus doichangensis seedlings were studied. In addition, the effects of abscisic acid (ABA) and salicylic acid (SA) pretreatments on the tolerance of this species to temperature extremes were tested. The results show that the content of soluble sugars increased with the duration of acclimation to cold (4 °C), and the relative electrical conductivity and malondialdehyde content increased significantly after 7 d; however, the content of proline did not vary significantly. After acclimation to cold for 3 and 7 d, the semilethal low temperature (LLT₅₀) was 0.8 and 1.1 °C lower, respectively, compared with that of the control. The maximum quantum yield of photosystem II (measured as variable to maximum fluorescence ratio, F_v/F_m) decreased significantly after freezing treatments (-4 to -8 °C), however, less when the plants were pretreated with 1-100 mg dm^-3 ABA. Acclimation to heat did not increase the semilethal high temperature (LHT₅₀). A low concentration (1 mg dm^-3) of SA increased LHT₅₀, but medium and high concentrations (10 and 100 mg dm^-3) decreased it. F_v/F_m decreased significantly after a heat shock (45-54 °C). The pretreatment with 1-50 mg dm^-3 SA ameliorated a subsequent heat (48 °C) stress.

Pepper (Capsicum annum L.) fruit colours vary from green, ivory, or yellow at the juvenile stage to red, orange, and yellow at the mature stage. Carotenoid accumulation causes fruit colour formation in ripe Capsicum fruits, and capsanthin is a main carotenoid in ripe red pepper fruits. In this study, pepper cultivars with red, yellow, and orange fruits were employed and four key genes named phytoene synthase (Psy), lycopene-β-cyclase (Lcyb), β-carotene hydroxylase (Crtz), and capsanthin/capsorubin synthase (Ccs) in the capsanthin biosynthetic pathway were analyzed. Results show that a clear expression of the Ccs gene in cv. CK7 (yellow) was detected, but the expressions of the Psy and Lcyb genes were low comparing with the other tested cultivars. In addition, the expression of the Ccs gene was not observed in cv. R37-1 (orange). Furthermore, though the four key genes (Psy, Lcyb, Crtz, and Ccs) were detected in red cultivar R15, the expressions of them were low. Sequence analysis further indicated that the Ccs genes in cv. CK7 and cv. R37-1 had a high sequence identity (97.36 %). Interestingly, amino acid sequences coded by the Ccs genes from cv. CK7, cv. R37-1, and cv. R15 were the same. The above results suggest that the genes Psy, Lcyb, Crtz, and Ccs were required for capsanthin synthesis in pepper, and their varied expression patterns resulted in the variety of pepper fruit colours.

The present study was aimed at understanding the effects of long term supplemental UV-B (3.6 kJ m^-2 d^-1) on biomass production, accumulation of reactive oxygen species, lipid peroxidation, and enzymatic antioxidants in leaves and roots of Withania somnifera (an indigenous medicinal plant). Under the UV-B treatment, a reduction in biomass and an increased malondialdehyde content (a characteristic of lipid peroxidation) were observed in both the shoots and roots. Amongst ROS, H₂O₂ content increased under UV-B in the leaves, whereas it decreased in the roots, and superoxide radical production rate decreased in both the plant parts. The activities of all enzymatic antioxidants tested (ascorbate peroxidase, catalase, glutathione reductase, peroxidase, polyphenol oxidase, and superoxide dismutase) increased under the UV-B treatment, the increase being greater in the roots.

MYB transcription factors (TFs) are known to have important functions in regulating the biosynthesis of secondary metabolites in plants. In this study, LeMYB1, a member of the MYB gene family of Lithospermum erythrorhizon, was cloned via the rapid amplification of cDNA ends. The alignment of the predicted translations of LeMYB1 with other MYB proteins revealed that LeMYB1 contained an N-terminal R2R3 repeat and a high degree of amino acid identity to NtMYBJS1 which is involved in jasmonic acid signalling and phenylpropanoid biosynthetic pathway regulation. To determine the expression pattern of LeMYB1, its promoter was cloned and the sequence analysis was performed. The results revealed a number of potential regulatory motifs related to tissue-specific gene expression and abiotic and biotic stress responses. Real-time PCR results suggest that LeMYB1 was induced transiently during the early stage when L. erythrorhizon cells were transferred from a B5 growth medium to a M9 production medium for shikonin formation. Exogenous methyl jasmonate (MeJA), an effective inducer of shikonin biosynthesis, induced the rapid LeMYB1 expression. In contrast, a treatment with ibuprofen (IBU), an inhibitor of jasmonate biosynthesis, significantly inhibited the LeMYB1 expression. Another inhibitor of shikonin formation, 2,4-dichlorophenoxyacetic acid (2,4-D), also markedly repressed the expression of LeMYB1. Tissue-specific expression analysis showed that LeMYB1 mRNA was predominantly accumulated in roots where shikonin was synthesized. Thus, the LeMYB1 gene may be a valuable member of the R2R3-MYB family in L. erythrorhizon and is possibly involved in the regulation of shikonin biosynthesis.

In plant cells, anion channels and transporters are essential for key functions. Members of the chloride channel (CLC) family located in intracellular organelles are required for anion accumulation, pH adjustment, and salt tolerance. Here, we cloned a maize (Zea mays L.) CLC gene, named ZmCLC-d, and found that its transcription was up-regulated under cold, drought, salt, and heat stresses, and after hydrogen peroxide (H₂O₂) and abscisic acid (ABA) treatments. The overexpression of ZmCLC-d in Arabidopsis conferred tolerance to cold, drought, and salt stresses; this tolerance was primarily displayed by an increased germination rate, root length, plant survival rate, antioxidant enzyme (catalase, peroxidase, and superoxide dismutase) activities, and a reduced accumulation of Cl^- in transgenic plants as compared with wild type (WT) plants. The accumulation of H₂O₂ and superoxide anion in leaves of the ZmCLC-d-overexpressing plants is much less than that of the WT plants. The expressions of some stress related genes, such as CBF1, CBF2, CBF3, DREB2A, and RCI2A, increased to a greater extent in the ZmCLC-d-overexpressing plants than in the WT. Our results strongly suggest that ZmCLC-d played an important role in stress tolerance.

We compared photosynthetic performance between cucumber (Cucumis sativus L.) leaves acclimated to saturating irradiances with high red: far red (R:FR = 10) and normal R:FR (= 1.4) ratios. The net photosynthetic rate (P_N) and stomatal conductance (g_s) of the leaves acclimated to high R:FR were greater than those of the leaves acclimated to normal R:FR; the greater g_s partly explains the greater P_N. The greater g_s of the high-R:FR-leaves probably resulted from a higher stomatal density and/or a greater size. P_N of the high R:FR leaves was still greater than that of the normal R:FR leaves at the same intercellular CO₂ concentration (c_i). This indicates that non-stomatal factors also increased the photosynthetic capacity of the high R:FR leaves. The maximum Rubisco carboxylase activity estimated from a P_N-c_i curve analysis was also greater in the high R:FR leaves, however, the intrinsic water-use efficiency (WUE_i = P_N/g_s) of the high R:FR leaves was lower than that of the normal R:FR leaves.

Roles of an altered porphyrin biosynthesis and antioxidants in protection against chilling and heat stresses were evaluated in rice (Oryza sativa L.). When exposed to the same exposure time (6 or 30 h), heat-stressed (45 °C) plants exhibited a less oxidative stress as indicated by a lower dehydration, ion leakage, and H₂O₂ production compared to chilling-stressed (4 °C) plants. Malondialdehyde production also increased after a mild chilling stress, whereas it increased only after a long-term heat stress. The content of protoporphyrin IX, Mg-protoporphyrin IX and its methyl ester, and protochlorophyllide drastically declined under both the stresses, particularly under the long-term heat stress. Greater increases in catalase and peroxidase activities in heat-stressed plants indicate more cofactors supplied for hemoproteins compared to those of chilling-stressed and untreated control plants. Intermediates of carotenoid biosynthesis, zeaxanthin and antheraxanthin, also increased under the chilling and heat stresses. In comparison to chilling-stressed plants, heat-stressed plants were more efficient in porphyrin scavenging and antioxidant enzyme responses, which may play crucial roles in plant protection under temperature stress, thereby suffering less from oxidative stress.

The aim of this study was to investigate the effects of salinity on germination, seedling growth, free polyamines (putrescine, spermidine, and spermine), and ethylene metabolism of two species (Atriplex prostrata Bouchér and Plantago coronopus L.) with different salt sensitivities. Seeds collected from Barranco Hondo (salt marshes, Jaén, southern Spain) were germinated at 0, 50, 100, and 200 mM NaCl in a growth chamber. The germination of P. coronopus seeds decreased considerably with an increasing NaCl concentration, however, seeds of A. prostrata showed high germination percentages (84, 87, and 80 %) at 0 (control), 50, and 100 mM NaCl, respectively, and only at 200 mM NaCl, the germination was reduced to 25 %. In the early phase of vegetative growth (8-d-old seedlings), the fresh mass increased in A. prostrata at 50 and 100 mM NaCl but the fresh mass of P. coronopus showed no significant differences. With respect to polyamines, there was a decrease of the putrescine and spermidine content at all the NaCl treatments, however, the spermine content increased and was much higher in P. coronopus than in A. prostrata. The ethylene, 1-aminocyclopropane-1-carboxylic acid content, and the 1-amino-cyclopropane-1-carboxylic acid synthase activity increased with the increasing NaCl concentration in A. prostrata, and only the ethylene content in P. coronopus. These results indicate that P. coronopus increased the free spermine content, whereas A. prostrata increased the ethylene biosynthetic pathway in order to survive in the saline conditions.

The role of chilling in bud dormancy release and biochemical changes in different organs were evaluated in stem cuttings of pear (Pyrus pyrifolia) cv. Cuiguan selected at the leaf fall. The cuttings were exposed to 5 °C for 0, 100, 200, 300, 400, 500, 600, and 700 chilling hours (named positive chill units; PCU). A 50 % bud break was observed in floral and vegetative bud cuttings at 300 and 600 PCU, respectively. A mean time to bud break was inversely proportional to the chilling treatment. The low-temperature stimulated starch hydrolysis accompanied with sucrose accumulation in all organs. Sucrose and sorbitol content increased substantially peaking at 100, 400, and 100 PCU in floral buds, vegetative buds, and bark, respectively, thereafter decreased when buds approached chilling satisfaction (300 and 600 PCU for the floral and vegetative buds, respectively), and then increased again up to 700 PCU. Hexoses (glucose and fructose) accumulated constantly in the buds from 0 to 700 PCU. In bark, glucose and fructose content increased up to 400 PCU, and then gradually decreased. Total amylolytic and α-amylase activities increased in all organs, especially in the floral and vegetative buds up to 100 PCU and then decreased in the floral and vegetative buds before increasing again after endo-dormancy release. Invertase activity remained high in the buds during chilling satisfaction possibly because of translocation of sucrose to the buds which functioned as a strong sink. The results suggest that a low availability of hexoses may be the cause of limited bud breaks due to lack of chilling. Chilling satisfaction of the buds may increase the content of soluble sugars and acid invertase activity, and decrease the starch content, which may correlate with improved bud breaks.

In vitro induction of haustoria from Castilleja tenuiflora Benth. was achieved by applying 25 μM catechin, 25 μM vanillin, or 25 μM H₂O₂. Of the treatments tested, 25 μM vanillin was the strongest inducer of haustoria in C. tenuiflora roots in vitro (up to 3 haustoria per root). Haustorium development occurred laterally and was observable 14 d after inducer application. It was characterized by elongation of the epidermal cells and division of the inner cortical cells which also possessed abundant nuclei. Histochemical analysis using 3,3-diaminobenzidine (DAB) and diphenylboric acid 2-aminoethyl ester (DBPA) indicated that the formation of haustoria was associated with the accumulation of H₂O₂ and flavonoids.

Tomato (Solanum lycopersicum Mill.) is sensitive to chilling stress during all stages of plant development. Genetic variation for chilling tolerance exists between cultivated tomato and its related wild species, but intra-specific variation has not been thoroughly investigated so far. Seedlings of 63 tomato accessions were evaluated under low temperature and two contrasting cultivars were identified for the trait: Albenga and San Marzano, the former being more chillingtolerant. To clarify the molecular mechanisms of chilling tolerance in tomato, changes in candidate gene expressions in the two tomato genotypes were analysed, using quantitative RT-PCR. Candidate genes were chosen among those known to be induced by chilling and/or with putative roles in CBF/DREB and ROS-mediated pathways. Results show that besides a CBF regulon, whose function is conserved, ROS and C2H2-type zinc finger protein-mediated cold signalling pathways were also involved in chilling tolerance. Under the chilling stress, the up-regulation of respective transcripts was consistently higher in the chilling-tolerant genotype than in the chilling-sensitive ones.

The effect of a rare earth element europium (Eu) on the long-distance transport of a plant defence compound quercetin (Q) was investigated. The complex Q/Eu³⁺ was synthesized in a HEPES buffer and tested for its transport ability through Arabidopsis thaliana and its ability to interact with target molecules in plant cells. Our results show that complexation with Eu³⁺ enhanced the transport of Q through Arabidopsis roots. During the transport, the complex degraded and released a free Q to tissues where Q was originally not available. Thus, the plant became better supplied with the defensive compound Q. The obtained spectrophotometric data imply that one of the reasons for the Q/Eu³⁺ degradation was the interaction of the complex with double stranded RNAs (dsRNAs) present in Arabidopsis. Since dsRNAs are replicative forms of plant RNA viruses, the ability of Q/Eu³⁺ to release a free Q in their presence suggests further investigation of this complex as a potential antiviral agent.

Innate immune system is employed by plants to defend against phytopathogenic microbes through specific perception of non-self molecules and subsequent initiation of resistance responses. Current researches elucidate that plants mostly rely on cell surface-located pattern recognition receptors (PRRs) and intracellular nucleotide-binding leucine-rich repeat proteins (NB-LRRs) to recognize pathogen-associated molecular patterns (PAMPs) and effector proteins from microbial pathogens, initiating PAMP- and effector-triggered immunity (PTI and ETI), respectively. Some pathogenic bacterial effector proteins are usually secreted into plant cells and play a virulence function by suppressing plant PTI, implying an evolutionary process of plant immunity from PTI to ETI. In the past several years, a great progress has been achieved to reveal fascinating molecular mechanisms underlying the pathogenic recognition, resistance signaling transduction, and plant immunity evolution. Here, we summarized the latest breakthroughs about these topics, and offered an integral understanding of plant molecular immunity.

A wheat stay-green mutant, named tasg1, was generated using the mutagen ethyl methane sulphonate applied to wheat (Triticum aestivum L.) cv. HS2. A drought stress was imposed by controlling irrigation and sheltering plants from rain. The antioxidant defence was characterized in the flag leaves of the tasg1 and wild-type (WT). Compared with WT, tasg1 had higher reduced ascorbate/oxidized ascorbate ratio, reduced glutathione/oxidized glutathione ratio, and antioxidant enzyme activities during senescence under both normal and drought stress conditions. The DHAR gene expression remained higher in tasg1 than in WT during the drought stress and tasg1 had a higher antioxidant defence competence which may contribute towards the delayed leaf senescence. The different transcriptional responses of some wheat senescence-associated genes to the drought stress between tasg1 and WT were observed. These results suggest that the competent antioxidative capacity might play an important role in the enhanced drought tolerance in tasg1.

Chloride channels (CLCs) play pivotal roles in plant development and anion transport. However, little research has been conducted about the CLC in fruit-bearing plants. Here we provide an insight into the evolution and expression patterns of CLC gene family members in various tissues of trifoliate orange [Poncirus trifoliata (L.) Raf.] and their responses to several treatments. Genome-wide analysis identified six PtrCLC genes. The predicted proteins had similar numbers of amino acids, but shared a low sequence identity. Phylogenetic analysis revealed that PtrCLC were classified into two separate subgroups, and PtrCLC4 and PtrCLC6 in subgroup II were more closely related to bacterial CLCs. Sequence comparison with EcCLCA from Escherichia coli reveals that PtrCLC showed amino acid divergence in anion selectivity of CLC proteins. Real time qPCR analysis shows that PtrCLC genes, particularly PtrCLC6, preferentially expressed in leaves. Nitrogen deficiency irreversibly inhibited expression of PtrCLC genes except for PtrCLC1. In contrast, NaCl stress profoundly induced expression of PtrCLC genes, particularly PtrCLC2 and PtrCLC4, both of which were also upregulated by ABA treatment. The results presented here provide a solid foundation for a future functional research on citrus CLC genes.

The aim of this work was to elucidate the molecular and biochemical mechanisms that control L-ascorbic acid (AsA) content variation in Myrciaria dubia. The AsA was quantified by high-performance liquid chromatography, gene expression by real-time quantitative PCR, and enzyme activities by spectrophotometric methods from leaves and immature fruits of two genotypes (Md-60,06 and Md-02,04) with pronounced (about 2 times) differences in the AsA content. In either genotype, the fruit peel had ∼ 1.5 times more AsA than the fruit pulp and ∼ 15.0 times more than the leaf. All tissues examined demonstrated the capability for AsA biosynthesis through the D-mannose/L-galactose pathway because mRNAs of the six key genes [GDP-D-mannose pyrophosphorylase (GMP), GDP-D-mannose-3',5'-epimerase (GME), GDP-L-galactose phosphorylase (GGP), L-galactose-1-phosphate phosphatase (GPP), L-galactose dehydrogenase (GDH), and L-galactono-1-4-lactone dehydrogenase (GLDH)] and catalytic activities of the corresponding enzymes (GMP, GDH, and GLDH) were detected. The differential expressions of genes and enzyme activities mostly correlated with the respective AsA content. Thus, the expression of several genes of the D-mannose/L-galactose pathway determined the AsA content variation in tissues of M. dubia.

Isoflavone synthase (IFS; CYP93C) plays a key role in the biosynthesis of phenolic secondary metabolites, isoflavonoids. These compounds, which are well-known for their benefits to human health and plant defence, are produced mostly in legumes. However, more than 200 of them have been described in 59 other plant families without any knowledge of their respective IFS orthologue genes (with the sole exception of sugar beet). In this study, we selected IFS from Pisum sativum L. (CYP93C18) for functional expression. CYP93C18 was isolated, cloned, and introduced into Arabidopsis thaliana. The presence of the gene was shown by Southern blot analysis and its expression in the transgenic Arabidopsis was proven by RT-PCR and Western blots. The functional activity of the heterologous IFS was verified by HPLC-MS analysis of the metabolite levels: the isoflavone genistein and its derivatives tectorigenin and biochanin A were detected in the overexpressing lines. In addition, 35S::CYP93C18::GFP fused proteins were transiently expressed in the leaves of Nicotiana benthamiana and the localization of the GFP signal was observed on the endoplasmic reticulum using confocal microscopy which is consistent with the data from the literature and with our in silico predictions. The putative mode of attachment of IFS to the endoplasmic reticulum membrane is suggested. The undemanding methodology presented in this paper is applicable to the functional analysis of newly-identified isoflavone synthase genes from various species.

An essential step in the carotenoid biosynthesis pathway is the formation of phytoene by phytoene synthase (PSY). In this study, three new genes coding PSYs (CpPSYA, CpPSYB, and CpPSYC) were cloned from Cucurbita pepo and their expression patterns analysed in three cultivars of summer squash which had a different carotenoid content. The gene sequences had a high similarity with those from other plant species, and their predicted proteins were significantly different from each other. A phylogenetic analysis indicates that CpPSYA and CpPSYB shared a high homology and were also homologous with PSYs from others cucurbits, whereas CpPSYC was more closely related to orthologues from strawberry and carrot. An expression analysis revealed that CpPSYA had a higher expression in flowers compared to leaves and showed a differential expression during fruit development. The amount of CpPSYA transcript was higher in fruits with a higher carotenoid content than in those with a lower carotenoid content. However, CpPSYB and CpPSYC showed a relatively high expression in leaves, and their expression in fruits varied among the different cultivars and fruit tissues. These results suggest that the CpPSY genes were under different regulatory mechanisms and they may have different roles in C. pepo.

The effects of foliar application of 24-epibrassinolide (EBR) on chlorophyll content, oxidative damage, and antioxidant capacity in eggplant (Solanum melongena L.) seedlings during a short-term chilling stress and subsequent recovery were investigated. The eggplant seedlings pretreated with 0.1 μM EBR were exposed to 5 °C for 12 h in the dark and then recovered under a normal temperature for another 12 h. The results show that EBR alleviated a chilling-induced oxidative stress reflected by a decrease in malondialdehyde and H₂O₂ content, O₂ ^.- production rate, and an increase in chlorophyll content and activities of superoxide dismutase, guaicol peroxidase, and catalase. The EBR pretreatment also promoted activities of ascorbate peroxidase, glutathione reductase, dehydroascorbate reductase, as well as the accumulation of ascorbic acid (AsA) and glutathione (GSH) involved in the AsA-GSH cycle. Moreover, after the12 h recovery, EBR plus chilling treated leaves showed no significant changes in content of MDA and chlorophyll a, lower reactive oxygen species (ROS) content, and greater activities of AsA-GSH cycle enzymes than in seedlings only chilled. The results suggest that EBR alleviated an oxidative damage caused by the dark chilling stress and accelerated a recovery rate mainly through increasing the ROS scavenging system including the AsA-GSH cycle.

Roles for nitric oxide (NO) and reactive oxygen species (ROS) during pollen-tube growth have been well established in angiosperms, but there remains lack of information regarding their potential signalling roles in pollen tubes in gymnosperms. Here, the pollen-tube elongation of Arizona cypress (Cupressus arizonica Greene) was investigated. Nitric oxide, ROS, and actin were detected using their respective fluorescent probes. Both NO and ROS were observed in the nuclei of generative cells and pollen-tube cells, and in the cytoplasm in the tip region. An intracellular NO content in the pollen cells was lowered using an NO scavenger or an NO-synthase inhibitor. Similarly, an endogenous ROS content in the pollen cells was lowered using an NAD(P)H oxidase inhibitor. These treatments reduced pollen germination and pollen-tube growth, and induced severe morphological abnormalities. Inhibition of NO and ROS accumulation also severely disrupted the actin cytoskeleton in the pollen tubes. These data indicate that NO and ROS had signalling roles in pollen germination and pollen-tube formation in cypress.

To identify genes involved in the susceptibility to powdery mildew in wheat, genechip hybridization experiments were performed applying interactions between resistant and susceptible wheat cultivars and powdery mildew. Ten genes were found to express exclusively in the susceptible host after inoculation. The single cell transient expression experiments revealed that the RNA interference (RNAi) of two expressed sequence tag (EST) sequences (accession No. BQ168479 and CA648596) could enhance host resistance by 22 and 15 %, respectively. Full-length cDNA cloning revealed that they represented long non-coding RNAs (lncRNAs) and markedly accumulated during the powdery mildew primary infection.

The ATP binding cassette B/multidrug-resistance/P-glycoprotein (ABCB/MDR/PGP) subfamily is a member of the ABC protein family. Significant progress has been made in the functional characterization of ABCB genes, particularly in Arabidopsis thaliana. This review evaluates recent advances concerning the plant ABCB subfamilies including their evolution and structure, the involvement and regulation of ABCB-mediated auxin transport, and the roles of ABCBs in plant growth and development. Insights into specific functions of members of the ABCB subfamily and their mediation of various regulatory pathways are also presented.

The main task of annually grown fibrous roots is to supply aboveground organs with water and nutrients. The key for this function, the development and mortality of primary tissues during a root lifespan is still poorly understood, especially in plants grown under field conditions. The goal of this study was to investigate the differentiation and maturation of fibrous roots from the initial appearance until the senescence and death. We monitored the histogenesis and anatomy of first order fibrous roots of black cottonwood (Populus trichocarpa Torr. & Gray) sampled at a known age. The daily examination of the first order fibrous roots revealed that only primary tissues were differentiated during the first seven days after their initiation and were maintained during the root lifespan. We observed all stages of exodermis and endodermis formation regulation a fibrous root water and nutrients uptake capacity. A cytological analysis, examined if any cellular symptoms of autophagy could be detected in senescent roots, indicated that vacuolar cell death was involved in root mortality. Our results are the first report strongly suggesting that programmed cell death (PCD) could be engaged in the senescence of ephemeral fibrous roots. The anatomical results advance our understanding of how roots absorptive ability is related to precise timing of tissue development during primary growth and of fibrous root senescence after fulfilment of its tasks.

The present study exemplifies morphogenic roles played by copper and zinc during micropropagation of Rauvolfia serpentina, an important medicinal shrub. Incorporation of 20 μM CuSO₄ or 25 μM ZnSO₄ to a Murashige and Skoog (MS) medium with optimized concentrations of auxins and cytokinins induced a maximum number of shoots per explant (40.67 ± 1.76 and 45.47 ± 0.24, respectively). However, higher concentrations of both the micronutrients negatively affected the morphogenic potential. The pigment content of the regenerants increased up to the optimal concentrations of both metals and thereafter decreased, whereas the maximum proline content was at the highest concentrations used. In vitro rooting of healthy shoots was accomplished using 0.5 μM IBA in a half strength liquid MS medium with 8.20 ± 0.37 roots, and root length of 5.50 ± 0.14 cm per microshoot. The plants survived a hardening procedure and were successfully acclimatized to field conditions with 95 % survival.