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The sex-related spatial heterogeneity of gas exchange rates over the leaf surface under salt stress was investigated in the dioecious species, Populus cathayana Rehd. Cuttings were subjected to two salt regimes: 0 and 75 mM NaCl added to the Hoagland solution, the control and the treatment group, respectively. Measurements of gas exchange parameters were taken from over 40 sites on the surfaces of representative ‘non-stressed’ and ‘salttreated’ leaves which had the same insertion point for two sexual cuttings. Compared to the control group, the treatment group showed a significant decrease in the mean values of the following: water use efficiency (WUE), Chlorophyll a (Chl a) concentration, chlorophyll b (Chl b) concentration, concentration of carotenoids (Caro), total chlorophyll concentration (TC) in two sexes, and net photosynthesis rate (Pn), stomatal conductance (gs), and stomatal length/width ratio (SR) in females. Also, in the treatment group, females exhibited lower WUE, Pn, gs, E, Chl a, Chl b, TC, and SR than males. Comparison of contour maps showed that the net photosynthesis rate decreased gradually from apical to basal zones over the leaf surface occurred in the two sexes under natural conditions, but under salt stress, the opposite trend was found in females only. The results suggest that the heterogeneity pattern of the gas exchange parameters in response to salt stress between the two sexes is quite different due to different strategies employed by males and females to maintain the photosynthesis rate under salt stress. This heterogeneity phenomenon under salt stress may mainly be attributed to the chlorophyll pigments in males and the stomatal apertures in females.
In order to provide information for the development of molecular selection markers for drought tolerance improvement, the methods of prometric analysis, quantitative real-time PCR and field evaluation were employed for the identification of the differential expression of candidate genes under drought stress in maize. At seventeen, twenty-four and forty-eight hours of polyethylene glycol-simulated drought stress at the seventh leaf stage, leaf samples were collected from two drought-tolerant inbred lines for prometric analysis by two-dimensional electrophoresis and peptide mass fingerprinting. Fifty-eight proteins out of more than 500 were found in response to drought stress. Three drought-induced spots 2506, 3507 and 4506 showed sequence similarity with cinnamyl alcohol dehydrogenase, cytochrome protein 96A 8 and S-adenosyl-L-methionine synthase, respectively. The expression of two key enzymes to lignin biosynthesis was quantified by quantitative real-time PCR among three drought-tolerant and one drought-sensitive inbred lines under drought stress and well-watered control conditions. After a decrease at the beginning of drought stress, the expression of cinnamyl alcohol dehydrogenase and caffeate O-methyltransferase recovered at twenty-four hours of the drought stress in the three drought-tolerant lines, but not in the drought-sensitive lines. Leaf lignin content, anthesis-silking interval and grain weight per plant were investigated with six inbred lines of varying drought tolerance under drought stress and well-watered control. Drought tolerance coefficients of these three characters were calculated and the correlation coefficients among these drought tolerance coefficients were estimated. Significant difference in leaf lignin content was found among the inbred lines and in response to drought stress. Close correlations were observed between the drought tolerant coefficients for leaf lignin content and grain weight per plant, and between the drought tolerant coefficients for leaf lignin content and anthesis-silking interval. These results indicate that leaf lignin content is a useful index for evaluation of drought tolerance in maize. Molecular selection markers can be developed on the basis of differential expression of the candidate genes and applied to maize improvement for drought tolerance.
Plant cell walls primarily comprise lignin, which performs functions of mechanical support, water transport, and stress responses. Lignin biosynthesis pathway proceeds through metabolic grid featuring complexity and diversity in enzymatic reaction. Cinnamate-4-hydroxylase (C4H, EC 1.14.13.11) is the gene encoding enzyme that catalyzes the second step of phenylpropanoid pathway responsible for biosynthesis of lignin. A full-length cDNA of C4H (designated as GbC4H), which spanned 1816-bp with a 1518-bp open reading frame encoding a 505-amino-acid protein, was cloned from Ginkgo biloba. A GbC4H genomic DNA fragment, spanning 3249-bp, was cloned and found to contain two exons and one intron. GbC4H protein showed high similarities with other plant C4Hs to include conserved domains of cytochrome P450 family. GT-1, W-box, and Myb/Myc recognition sites involved in stress response were detected in a 1265-bp upstream promoter region of GbC4H. Phylogenetic analysis suggested the common evolutionary ancestor shared by plant C4Hs including the gymnosperm enzyme. pET-28a-GbC4H plasmid was constructed and expressed in Escherichia coli strain BL21. Enzymatic assay revealed that recombinant GbC4H protein catalyzes conversion of trans-cinnamic acid to p-coumaric acid. Expression analyses in different organs showed high expression of GbC4H in stems and roots, whereas low expressions was found in fruits, carpopodium, and petioles. Further analysis indicated linear correlation of lignin contents with transcript levels of GbC4H among different tissues. GbC4H transcription was increased by treatments with UV-B, cold, salicylic acid, and abscisic acid, indicating the possible role of GbC4H in response to stresses and hormonal signal. Understanding of GbC4H function could benefit molecular breeding and reinforcement of defense mechanisms in Ginkgo.
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