The similarity or differences of peroxidase isozymes in rootstocks and scions may influence their graft compatibility. This study was conducted to identify peroxidase isozymes that may be used as markers to predict compatibility between pear (Pyrus communis L.) and various quince (Cydonia oblonga Mill.) clones. `Bartlett' (BT) and `Beurre Hardy' (BH) pear cultivars are known to form incompatible and compatible grafts, respectively, with quince rootstocks. The two pear scion cultivars were budded on `quince A' (QA), `quince BA-29', and 15 selected quince clones from Turkey. Bark and cambial tissues were taken from nonbudded rootstocks and scions, and 4 cm above and below the graft union for peroxidase isozyme analysis performed by starch gel electrophoresis. Isoperoxidase analyses were also performed on samples from the graft unions collected 12 months after grafting. Many isozyme bands were observed commonly in the two scions; however, one anodal peroxidase A was detected in BH (compatible scion) but not in BT (incompatible scion) samples. This isoperoxidase was also detected in QA, Quince BA-29, and nine of the Turkish quince clones. Another isoperoxidase, band B, was detected in BH but not in BT or any of the rootstocks. However, the compatible (BH/QA) and moderately compatible (BT/BA-29) graft union tissues contained bands A and B whereas incompatible graft union tissues (BT/QA) lacked both. Graft union samples involving BT and five Turkish quince clones (705, 609-2, 702, 804, and 806) had both `A' and `B' isoperoxidases while one or both of these bands were absent in nonbudded graft partners. Field observations of 3.5 year-old grafts of BT and Turkish quince clones revealed that the vegetative growth (vigor) of BT scion was significantly greater, when grafted on these five clones, than that in graft combinations with other clones. We suggest that matching of isoperoxidase `A' in quince rootstocks and BH pear scion may be associated with a compatible graft combination. Additionally, presence of isoperoxidases `A' and `B' in the graft union tissues may be used as an indicator to predict a compatible graft between BT and quince rootstocks.
Hatice Gulen, Rajeev Arora, Ali Kuden, Stephen L. Krebs, and Joseph Postman
Yueju Wang, Michael Wisniewski, Richard Meilan, Minggang Cui, Robert Webb, and Leslie Fuchigami
Ascorbate peroxidase (APX) plays an important role in the metabolism of hydrogen peroxide in higher plants, affording them protection against oxidative stress. We studied the effect of overexpressing a cytosolic ascorbate peroxidase (cAPX) gene—derived from pea (Pisum sativum L.)—in transgenic tomato plants (Lycopersicon esculentum L.). Transformants were selected in vitro using kanamycin resistance and confirmed by polymerase chain reaction (PCR) and northern analyses. An APX native-gel assay indicated that, in the absence of stress, APX activity in transgenic plants was several times greater than that measured in wild-type (WT) plants. Several independently transformed lines were propagated and evaluated for resistance to chilling and salt stress. After placing seeds at 9 °C for 5 weeks, percent germination was greater for seeds obtained from transgenic lines (26% to 37%) compared to the WT (3%). Plants from transgenic lines also had lower electrolyte leakage (20% to 23%) than WT (44%) after exposure to 4 °C. Visual assessment of transgenic and WT lines exposed to salinity stress (200 or 250 mm) confirmed that overexpression of APX minimized leaf damage. Moreover, APX activity was nearly 25- and 10-fold higher in the leaves of transgenic plants in response to chilling and salt stresses, respectively. Our results substantiate that increased levels of APX activity brought about by overexpression of a cytosolic APX gene may play an important role in ameliorating oxidative injury induced by chilling and salt stress.
Steven F. Vaughn
Localization of enzymes in specific plant tissues is crucial to understanding their role in processes such as differentiation and disease resistance. The oxidative enzymes lipoxygenase (LOX; EC 188.8.131.52), peroxidase (PER, EC 184.108.40.206) and polyphenol oxidase (PPO; EC 220.127.116.11) have all been implicated as playing critical roles in plant disease resistance. The histochemical localization of all three enzymes in potato tuber slices was accomplished either directly on the tissue slices (for LOX) or by blotting of the tissue onto nitrocellulose membranes (for PER and PPO). LOX was visualized in specific tissues by the oxidation of KI to I2 via lipid peroxides and the subsequent reaction of I2 and endogenous starch to form a colored, insoluble complex. PER and PPO activities were visualized with 4-methoxy-α-naphthol and 3,4-dihydroxy-phenylalanine, respectively. Fractionation of the slices and determination of enzyme activities in the fractions confirmed the reliability of these techniques.
Nadeem A. Abbasi and Mosbah M. Kushad
Peroxidase (POD), polyphenoloxidase (PPO), and superoxide dismutase (SOD) activities were measured during several phases of bud development, ripening, and storage of `Oregon Spur Red Delicious' (Malus domestica Borkh) apple fruit. POD, PPO, and SOD activities were significantly altered as a result of bud development. At the early stages of fruit development, the activities of these enzymes were significantly higher, then declined as the fruit increased in size. The activities of these enzymes declined as the fruit reached full maturity. Fruits harvested 3 weeks before commercial maturity and stored for up to 6 months have higher enzyme activities than fruits harvested at or beyond commercial maturity. The activities of these enzymes in relation to fruit firmness, soluble solids, and ethylene will be determined and related to overall fruit quality.
Chien Yi Wang
The activities of catalase and superoxide dismutase decreased while peroxidase activity increased in zucchini squash (Cucurbita pepo L., cv. `Elite') during storage at 5°C. Preconditioning of squash at 15°C for 2 days prior to the cold storage reduced the decline of catalase activity and suppressed the increase in peroxidase activity. The superoxide dismutase activity remained higher in temperature conditioned squash than in untreated squash. These results indicate that acclimation to chilling temperature in squash may also involve modifications in the activities of catalase, peroxidase, and superoxide dismutase.
Su-Jeong Kim*, Chun-Woo Nam, Dong-Lim Yoo, Seung-Yeol Ryu, and Ki-Sun Kim
Iris hollandica `Blue Magic' was treated with deionazed water as a control, 3% sucrose (Suc), 3% sucrose plus 0.4 mm silver thiosulphate (Suc+STS), 3% sucrose plus 200 mg·L-1 8-hydroxyquinoline sulphate (Suc+HQS) and 3% sucrose plus 100 mg·L-1 benzyl amino-purine (Suc+BA) for 4hrs and then transferred to tap water. The vase life treated with Suc+BA was extended 4 days longer than that of control. The treatment Suc+STS or Suc+HQS did not improve vase life. The amounts of water uptake and transpiration by all treatments decreased after harvest, but those values were higher in cut iris treated with Suc+BA than in those with control. Cut flowers treated with by Suc+BA markedly improved water balance, comparing with control which was quickly changed to minus value. Anthocyanin content in petals of cut flower treated with Suc+BA was 3.5 fold higher than that of control. The treatment by Suc+BA delayed discoloration in petals and senescence of cut Iris. Peroxidase (POD) activities of all treatments were reached maximum at 4th day after treatment and decreased thereafter. POD activity was highest when the cut iris was treated with Suc+BA. These results show that the use of Suc+BA is most effective treatment for improving the vase life and quality of cut Iris flowers.
Madhurababu Kunta, H. Sonia del Rio, and Eliezer Louzada
Reactive oxygen species (ROS) are continuously produced during the normal aerobic metabolism and also under environmental stress conditions. They are the major damaging factors to the photosynthetic machinery under stress conditions and need to be scavenged for the normal growth of the plant. Ascorbate peroxidase (APX) is the key enzyme in detoxifying H2O2, one of ROS from chloroplast and cytosol. A cDNA encoding a putative APXcit was isolated from mature `Dancy' tangerine (Citrus reticulata Blanco) juice vesicles using differential display reverse transcription-polymerase chain reaction (RT-PCR). Subsequently, full-length APXcit cDNA clone and genomic clone were obtained and sequenced. The full-length APXcit sequence is composed by 1082-bp nucleotides, including an open reading frame (ORF) of 753 bp, encoding a protein of 250 amino acids (27 kDa). The 5' un-translated region (UTR) of the APXcit gene consisted of 91 nucleotides and the 3' UTR consisted of 238 nucleotides. Homology search for APXcit at GenBank database showed high similarity to APX from several plant species.
L. Mark Lagrimini, Jill Vaughn, W. Alan Erb, and Sally A. Miller
Lignin composition in leaf, fruit, and fruit outer epidermis of transgenic tomato (Lycopersicon esculentum Mill.) plants that overproduce the enzyme tobacco anionic peroxidase (TobAnPOD) was analyzed. This enzyme may catalyze the polymerization of cinnamyl alcohols into lignin in tobacco (Nicotiana tabacum L.); therefore, we predicted that its presence in the transformed tissue would increase lignin levels in healthy and wounded tissue. Lignin levels in healthy plants increased by 20% in leaf, 49% in fruit, and 106% in fruit outer epidermal tissue. Mature-green fruit were aseptically wounded and incubated in darkness for up to 7 days. Soluble phenols in wounded transgenic fruit increased by more than 300% hut changed little in control fruit. As with soluble phenols, lignin content in wounded transformed fruit increased by more than 20-fold hut increased less than two-fold in control fruit. Transgenic seedlings overproducing TobAnPOD were screened for susceptibility to several pathogens, but resistance did not increase. Possible TobAnPOD roles in lignin biosynthesis, phenol metabolism, stress response, and disease resistance are discussed.
Y.H. Huang, G.A. Lang, C.E. Johnson, and M.D. Sundberg
Five-year old `Sharpblue' southern highbush blueberry plants (Vaccinium corymbosum L.) were self- and cross-pollinated (`O'Neal') to study peroxidase (POD) activity, isozyme patterns, and histological localization during fruit development. Cross-pollination resulted in larger and earlier-ripening fruit. Activities of soluble and bound POD were very high during fruit growth period I, with peaks at 10 and 20 days after self- and cross-pollination. Activity was much higher for cross-pollinated fruit. During fruit growth period II, POD activities were low in both pollination treatments. During ripening, soluble POD increased, then declined in both treatments. Bound POD activities increased during the color transition from blue to dark blue, with the increase greater in self-pollinated fruit. Banding patterns of soluble and bound POD isozymes and their histological localization varied by pollination treatment as well as fruit developmental stage. During fruit ripening, soluble POD activity appeared to be associated with color transition from light blue to blue, while bound POD activity appeared to be associated with color transition from blue to dark blue.
Cheng-lie Zhang, Paul H. Li, and Charles C. Shin
Twenty-day-old `Bush Blue Lake 47' common bean plants grown in a growth chamber at 25 days/22C night and a 12-hour photoperiod regime were foliar sprayed with 0.5% GLK-8903 including 0.05% Tween-20. After 24 hours of treatment, plants were chilled in a cold room (4C day/night, 12 hours of light). After 3 days of chilling, leaves of untreated controls were injured, as visually characterized by leaf wilting, whereas leaves of the GLK-8903-treated plants still retained turgor. During chilling, the activity of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) decreased. GLK-8903 treatment had no effect on SOD and POD activities; however, the CAT activity was reduced significantly after GLK-8903 treatment either at 25 or at 4C. During chilling, the content of malondialdehyde, a decomposition product of phospholipid peroxidation, increased in treated plants and untreated controls, with increased content significantly lower in the former compared with the latter. The GLK-8903 per se and total lipid extracted from GLK-8903-treated plants were able to reduce the linoleic acid oxidation in vitro. The mechanism by which GLK-8903 alleviates chilling injury in bean plants is discussed.