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- Author or Editor: Kourosh Vahdati x
The genetic structure and diversity of natural populations of Juglans regia L. in Iran were characterized using 11 microsatellite loci. A total of 105 individuals from seven populations were sampled. A high level of genetic diversity was observed within populations with the number of alleles per locus (A) ranging from three to 11 (average = 5.73), the proportion of polymorphic loci was 100%, and the expected heterozygosity ranged from 0.598 to 0.848 (average = 0.707). The proportion of genetic differentiation present among populations accounted for 12% of the total variation. Such considerable interpopulation differentiation detected in J. regia L. could have resulted from several factors, including restricted gene flow between populations. Significant departures from Hardy-Weinberg equilibrium were observed for WGA276, WGA32, and WGA321 loci. The deviations were primarily the result of the surplus of heterozygotes. The unweighted pair group method with arithmetic mean cluster analyses based on Nei's unbiased genetic distances separated the seven populations into two main groups.
Somatic embryos (SEs) can play important roles in genetic manipulation and breeding. They can be used as targets for induced mutagenesis, as material for cryopreservation and germplasm conservation, and for transformation or gene editing in support of plant improvement and proof of gene function. However, germination rates of walnut (Juglans regia) SEs are low, and the genetic stability of plantlets regenerated from them has not been explored. Here, we studied first the effects of gibberellic acid (GA3) and low temperature storage (LTS) on germination of walnut somatic embryos. Second, we assessed the genetic fidelity of plantlets regenerated from these SEs by comparing them to each other and to their cultivar of origin. Results showed that GA3 and LTS increased the walnut SE germination rate. The best rate was observed when SEs were subjected to LTS for 60 d followed by culture on a medium with either 1 or 3 mg·L−1 GA3 (56.6% and 46.6% germination respectively). Genetic stability was evaluated, using flow cytometry and 15 sets of ISSR primers. Flow cytometry indicated that all samples (i.e., regenerated and parental counterpart) showed the same peak. Amplified fragments of inter simple sequence repeats (ISSR) primers ranged in size from ≈200 to 1800 bp. All ISSR profiles of regenerants were monomorphic. Results did not show any genetic differences among plantlets regenerated from SEs or from their parental counterpart. Due to this apparent genetic stability, walnut SEs can be useful for genetic transformation and germplasm conservation.
In vitro rooting of three commercial cultivars of Persian walnut (Juglans regia L.), `Sunland', `Chandler', and `Vina', was examined using a two-phase rooting procedure: root induction in the dark on Murashige and Skoog (MS) medium with 15 μm IBA followed by root development in the light on a mixture of one-quarter strength Driver Kuniyuki Walnut (DKW) basal medium and vermiculite (1:1.25, v/v). Rooting percentages were: `Sunland' (94%), `Chandler (55%), and `Vina' (27%). A positive relationship was observed between the vigor of cultivars and rooting ability, but shoot length did not affect rooting success. Rooting was optimum when shoots were cultured on root induction media for 6 to 8 days. Increasing the sucrose level in the root induction medium to 40 g·L-1 improved rooting, and shoots induced to root at 22 °C rooted more readily than those induced at 30 °C. Either increasing or decreasing the nitrogen level in the multiplication medium had a negative effect on rooting. Rooted walnut shoots often cease growth during acclimatization, resulting in shoot rosetting. Spray application of Promalin® (25 mL·L-1) caused buds to break and induced elongation of shoots. Chemical name used: indole-3-butyric acid (IBA).
Walnut (J. regia L.) is one of the most sensitive plants to abiotic stresses for which finding salt-tolerant genetic resources is very important. Effects of salt stress on seed germination of seven walnut cultivars (Serr, Lara, Pedro, Chandler, Hartely, Vina, and Roun de Montignac) were studied. Salt stress treatments were applied using NaCl solutions ranging from 50 to 250 mm. The increase in salinity levels resulted in a substantial decrease in root relative water content. The percent of germination was also affected by salinity level and cultivar and their interactions. The mean germination time differed among treatment media and cultivars and a significant interaction was observed between these two factors. According to the cluster analysis, ‘Chandler’ was classified as the most tolerant cultivar. ‘Hartley’, ‘Pedro’, and ‘Round de Montignac’ (‘RDM’) were also classified in another group (semisensitive), whereas ‘Vina’, ‘Serr’, and ‘Lara’ were classified as sensitive cultivars. With the increase in salt stress, resistant cultivars accumulated more potassium (K) and calcium, especially in their shoots. However, in the semitolerant cultivars, accumulation of K in the root was more than in the shoot. Differences in magnesium accumulation in roots and shoots of all samples were significant at all stress levels and were dependent on genotypes. Sodium levels in roots were higher than in the shoots in most cultivars, especially in the semisensitive and tolerant ones. There was a low correlation (r 2 ≤ 0.15) between walnut seed traits (seed weight and kernel weight) and growth indices with respect to salinity tolerance.
The objective of this work was to determine the chilling and heat requirements of Persian walnut cultivars and genotypes using excised twigs. The experiment was carried out from Nov. 2006 and 2007 to Mar. 2007 and 2008. One-year-old twigs were prepared from four cultivars and four domestic genotypes of Juglans regia L. After leaf fall, the twigs were taken and placed in plastic bags and kept at 4 ± 1 °C to stimulate 400 to 1500 chilling hours. After chilling, the excised twigs were transferred to the greenhouse with a natural photoperiod and a temperature from 18 to 27 °C. The evaluation of budbreak was made three times a week and the number of accumulated growing degree hours (°C) was determined until the buds reached the balloon or green tip stage. The chilling requirements were lowest (400 h) for catkins and highest (1000 h) for lateral buds. The Serr cultivar and ‘Z30’ genotype had the lowest chilling requirements (650 and 650 h). ‘Lara’, ‘Z63’, ‘Z53’, ‘Pedro’, and ‘Z67’ showed intermediate chilling requirements with values of 900, 900, 800, 750, and 750 h, respectively. Finally, ‘Hartley’ completed its dormancy after an accumulation of 1000 h, being the walnut cultivar with the highest chilling requirement in our study. As the final result, the cultivars and genotypes were classified into three groups based on their heat requirements: low requirement (‘Z30’ and ‘Serr’), medium requirement (‘Z53’, ‘Z67’, ‘Lara’, and ‘Pedro’), and high requirement (‘Hartley’ and ‘Z63’).
Acclimatization of in vitro plantlets is one of the key steps in successful tissue culture propagation. Gaseous atmosphere during in vitro culture can influence the rate of ex vitro acclimation of the plantlets produced. In the current study, effects of elevated CO2 concentration on the leaf water loss dynamic responses of in vitro–produced walnut leaves during ex vitro desiccation were investigated. Elevated CO2 concentration in the headspace of culture vessels caused a considerable decrease in stomatal aperture. Although the traits related to stomatal size were not influenced by CO2 elevation, the number of small stomata was increased, and the number of large stomata was decreased at elevated CO2 concentration. Higher CO2 concentration resulted in a lower transpiration rate and a higher relative water content (RWC) during ex vitro desiccation. This improvement was due to decreased stomatal aperture during the first phase of water loss. Osmotic potential (ψs) was decreased under an elevated CO2 concentration, but no influence was observed on the concentration of compatible solutes. In conclusion, increasing the CO2 concentration of culture vessel headspace can be an efficient tool for improving acclimation of in vitro–grown walnuts without negative effects on plantlet growth.
To study the cold-hardiness of Persian walnut cultivars and selections, three methods were compared: 1) thermal analysis; 2) evaluation of tissue health after controlled freezing; and 3) field observations after a severe midwinter freeze. Stem segments and buds were collected from eight Persian walnut genotypes (four commercial cultivars and four promising Iranian selections). Thermal analysis was conducted using thermoelectric modules (TEM) to measure the high (HTE) and low (LTE) temperature exotherms produced when water and tissues freeze. TEM signals were recorded as the temperature of the samples was decreased at a rate of 2 °C/h. Tissue injury under controlled temperatures was evaluated using pre-chilled stem segments cooled at 2 °C/h to set temperatures ranging from –5 to –30 °C and then held at these temperatures for 16 h. Frozen samples were thawed and visually evaluated for severity of injury. Cold damage under field conditions was evaluated after an unusually severe winter freeze. Twigs from affected trees were removed in mid-February and in April and visually rated for extent of injury and ability to recover. The occurrence of LTEs was correlated with death of the tissues as assessed by tissue browning. Both the capacity to supercool and the cold-hardiness of cultivars and selections tested increased with accumulated seasonal chilling and decreased as they approached spring budbreak. Thermal analysis showed a tendency for buds and stems to exhibit multiple LTEs at peak dormancy. The cultivars and selections were classified into three groups based on their cold-hardiness: sensitive (‘Z30’ and ‘Serr’), semihardy (‘Z53’ and ‘Z67’), and hardy (‘Lara’, ‘Hartley’, ‘Z63’, and ‘Pedro’).
Correlation and causal relationships among 21 horticultural traits were determined using 71 walnut genotypes selected from seven valleys in Kerman Province, Iran. Pearson's correlation coefficient was calculated. Kernel percentage and blight susceptibility were used as dependent variables in a stepwise regression model to determine predictor variables. Direct and indirect effects of each independent variable were calculated using path analysis. A highly significant correlation was observed between lateral bearing habit and yield. Lateral-bearing trees were also more susceptible to blight and winter cold than terminal bearers. Kernel and nut weights, shell thickness, and difficulty of extracting kernel halves were the most important traits accounting for kernel variation. Kernel weight and difficulty extracting kernel halves had the strongest positive direct effects and nut weight the most negative. Flowering habit, nut shape, and leafing date had positive direct effects on blight susceptibility, but the large residual effects suggest there are other important determinant traits for blight susceptibility, which were not considered in this study.
An experiment was conducted in a persian walnut (Juglans regia) orchard in the north of Iran to evaluate the effects of zinc (Zn) as zinc sulfate and/or boron (B) as boric acid in foliar spray with different concentrations and combinations. Three B and three Zn concentrations (0, 174, and 348 mg·L−1 for B and 0, 1050, and 1750 mg·L−1 for Zn) were applied either independently or in combination. Leaf nutrient concentrations, pollen germination, fruit set, leaf chlorophyll index, nut and kernel characteristics, vegetative growth, nut weight, and nut yield were measured to assess the effects of treatments. The results showed that all B and Zn applications and combinations had a significant effect on all traits except nut and kernel diameter, shell percent, husk thickness, and pistillate flower abscission (PFA). Pollen germination, fruit set, vegetative growth, nut weight, kernel percent, nut and kernel length, and chlorophyll index were highest when B and Zn were applied simultaneously at 174 and 1050 mg·L−1 concentrations, respectively.
The effects of osmotic stress induced by polyethylene glycol on the seed germination of 16 walnut genotypes (‘Z30’, ‘Z53’, ‘Z67’, ‘Z60’, ‘Z63’, ‘K72’, ‘B21’, ‘V30’, ‘Panegine20’, ‘Hartley’, ‘Pedro’, ‘Vina’, ‘Lara’, ‘Serr’, ‘Ronde de Montignac’, and ‘Chandler’) of Juglans regia L. were studied. Potted seeds were kept under controlled conditions (12/12-h light/dark photoperiod and 25 ± 1 °C) during the experiments. The objective was to screen genotypes and determine the critical range of osmotic potential (ψS) for walnut seeds during germination. Decreasing the ψS of the germination solutions markedly reduced germination percentage in all genotypes, but there were variations in degree among the genotypes. The Z genotypes were the most sensitive to osmotic stress, and their germination rates were the lowest at ψS more negative than –0.75 MPa. Cluster analysis produced a dendrogram with four groups differing in their tolerance to osmotic stress. Based on factor analysis, four factors explained 90.45% of data total variance. Factor analysis showed that tissue fresh and dry weight, tissue water content, and thickness were the most important traits under drought condition. Regression analysis failed to show a significant relationship between percent germination and either seed weight (r 2 = 0.0601) or kernel weight (r 2 = 0.0258).