Hatice Gulen

Cold-hardiness of olive (Olea europaea L.) cultivars in cold-acclimated and non-acclimated stages: seasonal alteration of antioxidative enzymes and dehydrin-like proteins.

SUMMARY Seasonal patterns of antioxidative enzymes and proteins and their relations to cold-hardiness of nine olive (Olea europaea L.) cultivars (Ascolona, Domat, Gemlik, Hojoblanca, Lecquest, Manzanilla, Meski, Samanli and Uslu) are documented in the current study. Fully expanded, uniformly sized leaves from 2-year-old shoots of the cultivars were collected from 20-year-old trees in cold-acclimated (CA, in January) and non-acclimated (NA, in July) stages. Leaf samples were exposed to low temperature at 4, x5, x10 and x20 xC for 12 h to determine their cold-hardiness (LT50; assessed by electrolyte leakage). Cold-acclimation produced an increase in freezing tolerance of all cultivars (by lowering LT50). Domat and Lecquest were found to have the highest cold-hardiness among the nine cultivars investigated. Ascolona, Gemlik, Hojoblanca had moderate cold-hardiness, while Samanli, Meski, Uslu and Manzanilla were more sensitive. Activities of catalase (CAT : EC 1 . 11 . 1 . 6), ascorbate peroxidase (APX : EC 1 . 11 . 1 . 11) and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase significantly varied depending on the cold-acclimation stage and the cold-hardiness level of the cultivars. Activities of the three antioxidative enzymes and total soluble proteins (TSP) were higher in the CA stage than in the NA stage. Although no accumulation of major polypeptides, except a 23 kDa protein, was detected either in CA samples or NA samples by SDS-PAGE, anti-dehydrin immunoblots revealed that the 43 and 23 kDa polypeptides were detectable during cold-acclimation of olive cultivars. Accumulation of both 43 and 23 kDa dehydrin was significantly higher in the CA stage than in the NA stage in all cultivars. Accumulation of 43 kDa dehydrin was correlated with cold-hardiness of the cultivars, while 23 kDa dehydrin was considered as cultivar-dependent since its accumulation was not parallel to LT50 values of the cultivars. Indeed, the tissues of cvs Domat, Lecquest, Ascolona, Hojoblanca and Gemlik were found to enhance the structural stability of cellular membranes in the CA stage by increasing both the activity of such enzymes as CAT, APX and NADPH oxidase to activate the antioxidative systems and the expression of 43 kDa dehydrins.

Some physiological changes in strawberry (Fragaria × ananassa ‘Camarosa’) plants under heat stress

Summary The effects of heat injury induced by long exposures were evaluated in strawberry (Fragaria × ananassa ‘plants’) Camarosa in this study. Seedlings were grown in 14 × 12 cm pots using perlite for three weeks at 25/10°C day/night temperature, and watered daily by modified 1/3 Hoagland nutrient solution. Half of the plants were transferred to a growth chamber with a constant 25°C, 16/8 h (light/dark) photoperiod regime and 1200 lux light intensity for a week to acclimate the plants. Temperature was increased stepwise (5 K per 48 h) to 30, 35, 40°C and finally to 45°C. In addition to others, plants were transferred from the outside to the growth chamber, at each temperature step to impose a heat shock. Leaf relative water content (RWC, %), loss of turgidity, chlorophyll content (Spad value) and heat-stress tolerance (HTS; LT50) were measured in control and stressed plants. Total soluble proteins and total DNA were extracted from the leaves following the above treatments using standard procedures and total protein contents were determined using a Bradford assay. In general, effects of gradual heat stress (GHS) and shock heat stress (SHS) on the variables studied were mostly significant, except for chlorophyll content, while the effect of temperatures was significant for all the variables. Interaction between the heat stress type and temperature treatments was not significant for leaf RWC, loss of turgidity and chlorophyll content. Data also indicated that total protein and DNA contents were changed significantly by heat stress types (GHS and SHS) and/or temperature treatments. The plants exposed to GHS exhibited a significant increase in HST compared with the plants exposed to SHS (LT50 of 41.5°C and 39°C, respectively). Consequently, gradual heat stress increased HST in strawberry leaves. Increased HST may be associated with the accumulation of several heat-stable proteins in GHS plants.

Annual patterns of total soluble sugars and proteins related to coldhardiness in olive (Olea europaea L.‘Gemlik’)

Summary Annual patterns of cold hardiness, total soluble sugars (TSS) and proteins (TSP) were characterised during two consecutive years (May 2003–April 2005) in leaf and bark tissues of olive (Olea europaea L.) ‘Gemlik’ trees. One-year-old shoots from 13-year-old olive trees were collected each month and exposed to low temperature at 4°C, –5°C, –10°C, or –20°C for 12 h to determine their cold-hardiness (LT50; assessed by electrolyte leakage). A considerable increase in cold-hardiness was observed with decreasing temperature. Leaf and bark tissues exhibited an increase in cold-hardiness during the Autumn, which reached a maximum in mid-Winter, then gradually decreased during the Spring, and reached a minimum in mid-Summer. Results indicated that leaf and bark tissues in both experimental years responded similarly to cold, with a continuous increase in their TSS and TSP contents during field adaptation in Autumn and Winter, which paralleled their cold-hardiness. The annual cycle of TSP from leaf and bark tissue was characterised by one dimensional SDS-PAGE. A seasonal fluctuation was observed in leaf proteins of 66 kDa and 43 kDa, and in bark proteins of 70 kDa, 43 kDa and 16 kDa, which paralleled cold-acclimation. Evaluation of these leaf and bark TSP profiles showed that these polypeptides disappeared in the Summer, followed by their accumulation again during the Autumn and Winter. Moreover, all these polypeptides became less visible during the Spring. The relationships between these five proteins, TSS contents, and cold-hardiness in olive are discussed.

The activities of catalase and ascorbate peroxidase in olive (Olea europaea L. cv. Gemlik) under low temperature stress

In this study, one-year-old shoots of the olive (Olea europaea L.) cv. Gemlik were tested at artificial low temperatures (4, −5°C, −10°C, and −20°C) every month for two years. For low temperature treatment, the degree of cell membrane injury in leaves and barks was determined by ion leakage method. In addition, with regard to antioxidative defense mechanism, activities of catalase (CAT, EC 1.11.1.6) and ascorbate peroxidase (APX, EC 1.11.1.11) enzymes were determined. Leaf and bark tissues subjected to 4°C and −5°C injured to a limited extent in all months. However, more than 50% injury occurred by temperatures equal to or colder than −10°C treatments depending on the season. For −10°C and −20°C treatments, the lowest and the highest injury in leaf and bark tissues were detected during winter and summer seasons, respectively. We determined in this study that CAT and APX enzyme activities are generally higher during fall and winter compared with those in summer. On the other hand, CAT and APX enzyme activities started increasing during fall along with a decreasing freezing injury while the activities of these enzymes decreased to some extent during winter when freezing injury was the lowest. In addition, while CAT activity decreased with low temperature treatments, APX activity did not change until −5°C treatment but decreased with decreasing temperatures starting from −10°C depending on the month the tissue was obtained. In conclusion, olive plant shows considerable tolerance to low temperatures that are achieved after daily gradual decreases by increasing cell membrane stability through complicated mechanisms including antioxidative enzyme metabolisms. In addition, APX may be more effective in maintaining cold-hardiness of olive compared with CAT.

Determination of self-incompatibility groups of sweet cherry genotypes from Turkey.

Determination of S-allele combinations of sweet cherry genotypes and cultivars has importance for both growers and breeders. We determined S-allele combinations of 40 local Turkish sweet cherry genotypes using a PCR-based method. Ten different S-alleles were detected. Although the most common S-allele was S3, as also found in Western genotypes and cultivars, there were some differences in the frequencies of some S-alleles between Turkish and Western sweet cherry genotypes. According to their S-allele compositions, 30 local Turkish sweet cherry genotypes were assigned to 10 previously identified incompatibility groups. For the remaining genotypes, whose S-allele combinations did not fit to any previous incompatibility groups, three more incompatibility groups, XLII, XLIII and XLIV, were proposed. Results obtained from this study will help both sweet cherry growers and breeders to better manage these local Turkish sweet cherry genotypes in their orchards.

Assessment of genetic relationships among 29 introduced and 49 local sweet cherry accessions in Turkey using AFLP and SSR markers.

Summary The characterisation of sweet cherry (Prunus avium L.) genetic resources in Turkey may help to increase their use in breeding programmes worldwide, as Turkey is the centre of origin of sweet cherry. Amplified fragment length polymorphism (AFLP) and simple sequence repeat (SSR) markers were therefore used to analyse genetic diversity among a total of 78 local and introduced sweet cherry cultivars. Four AFLP primer combinations, and six SSR primer pairs for sweet cherry were used for genetic diversity analysis. A genetic similarity matrix was calculated using the combined data from AFLP and SSR analyses with simple matching coefficient. Genetic similarities among the sweet cherry genotypes studied were higher than 42%. No two accessions had an identical AFLP and SSR marker profile, indicating that all 78 genotypes were unique. An UPGMA dendrogram, based on the similarity matrix, revealed 18 separate Groups at or above the 70% similarity level. While some Groups consisted of both introduced and local genotypes, other Groups had only local genotypes. This result suggests that there was broad genetic diversity among the local Turkish sweet cherry genotypes, which was not present in the introduced sweet cherry accessions. The genetic variation present in local Turkish sweet cherry genotypes may be useful for future breeding programmes. We found that the use of both SSR and AFLP marker systems was effective for distinguishing between genetically-close sweet cherry genotypes. These marker systems can be used to complement pomological and morphological markers during the characterisation and identification of sweet cherry genotypes.

Cold hardiness of olive (Olea europaea L.) cultivars in cold-acclimated and non-acclimated stages: seasonal alteration of soluble sugars and phospholipids.

In many plant species, several physiological and biochemical changes occur during low-temperature-induced cold acclimation. A previous study with olive cultivars (Cansev et al. 2009) demonstrated a correlation between the level of accumulation of certain leaf proteins besides antioxidative enzyme activities and cold hardiness of the cultivars. The present paper analysed soluble sugar (SS) and phospholipid (PL) contents of cold-acclimated (CA) and non-acclimated (NA) leaf tissues in order to explain the mechanism of cultivar-dependent response to cold in olive. In general, cold acclimation significantly increased total soluble sugar (TSS), reducing sugars and sucrose contents of all cultivars to various extents depending on the cold hardiness of cultivars. In addition, TSS, reducing sugars and sucrose contents in cold-tolerant cultivars were significantly increased, whereas TSS, reducing sugars and sucrose contents in cold-sensitive cultivars either did not change or increased slightly in CA stage compared with those in NA stage. Even though reducing sugars were the major soluble sugar in olive leaves, levels of sucrose accumulations in CA stage compared with those in NA stage were greater than those observed in reducing sugars accumulation. Changes in levels of total PL, as well as the three individual PL fractions phosphatidylcholine (PC), phosphatidylethanolamine (PE) and phosphatidylinositol (PI), were investigated in olive leaf tissues. Significant increases in levels of PC and PE fractions during CA compared with those in NA stage suggested that PC and PE maintained the cold hardiness of olive cultivars more effectively than did PI. Although the precise mechanisms by which olive responds to cold may still be open to discussion, soluble sugars and PL are clearly important in the ability of olive cultivars to stand against cold stress.

Effect of processing techniques on antioxidative enzyme activities, antioxidant capacity, phenolic compounds, and fatty acids of table olives

In this study, olive fruits (Olea europeae cv. Gemlik) of the most common sources of table olives in Turkey were used. Total polyphenol content (TPC), antioxidant capacity (AC), and antioxidant enzymes (catalase, CAT; ascorbate peroxidase, APX; and glutathione reductase, GR) of table olives were compared by 4 different methods of ripe table olive processing. Results revealed that TPC of the processed olives ranged from 117.44 to 418.69 mg gallic acid equivalents/g fresh weight (f.w.). The highest AC as mg Trolox equivalents of 189.58/g f.w. was obtained from unprocessed black olives. CAT, APX, and GR activities of unprocessed olives were higher than those obtained in all processed olives. In conclusion, TPC, AC, and antioxidant enzyme activities are strongly affected by fruit ripening and processing in table olives of ‘Gemlik’ cultivar. In addition, the best processing technique is untreated black olives in brine for antioxidant properties.

Micropropagation of Some Vegetative Rootstocks of Sweet and Sour Cherry by Meristem Culture

This study was carried out to determine best culture media for the in vitro propagation of sweet and sour cherry rootstocks. Two different rootstocks which are called Edabriz and Damil were cultured in this study. Meristems showed better development when they are cultured in MS medium supplemented with GA3. 2 mg/1 BAP was used in the multiplication stage. Two-step rooting procedure was applied in the rooting stage; full MS medium and 0.5xMS medium supplemented with IBA (3 mg/1). Both of them were prepared solid and liquid and all the cultures were kept in the dark during one week. At the end of the week, they were transferred new rooting media which are include 1 mg/1 IBA (solid and liquid),Damil showed best root development in Bl solid medium (0.5xMS medium + 1 mg/1 IBA). Edabriz showed best root development in Al solid medium (MS medium + 1 mg/1 IBA). More hairy root formation was obtained from these media.