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’).
Asadolah Aslani Aslamarz, Kourosh Vahdati, Majid Rahemi and Darab Hassani
Mohammad Sadat Hosseini Grouh, Kourosh Vahdati, Mahmoud Lotfi, Darab Hassani and Nejat Pirvali Biranvand
We report the first successful regeneration of haploid lines in persian walnut (Juglans regia) developed by in situ parthenogenesis followed by embryo rescue. Female flowers of cultivars Hartley and Pedro and two native Iranian selections (Z63 and Z67) were pollinated using pollen of selections Z53 and Z30 that had been irradiated with gamma rays at five doses (50, 150, 300, 600, and 900 Gy). Gamma-irradiated pollen induced fruit set and development of some parthenogenetic embryos. The immature embryos were excised 30 and 45 days after pollination, cultured in vitro, and then stratified for 30 days at 4 °C to overcome dormancy. Ploidy level of the resulting plantlets was determined by chromosome counting and flow cytometry. Haploid plants were obtained from ‘Hartley’, ‘Pedro’, Z63, and Z67 after pollination using pollen irradiated at 300 and 600 Gy. Plants obtained from pollen irradiated at 50 and 150 Gy were all diploid. Molecular marker analysis using four simple sequence repeat (SSR) markers also showed that all the diploid plants recovered were zygotic and no spontaneous double haploid plants were obtained in this work. Also, the haploid plantlets presented only one allele of their female parents. These profiles confirmed the parthenogenetic origin of the obtained haploid plants. The techniques used to induce haploid walnut plants by irradiated pollen were successful and could be used in breeding programs and accelerate genome analysis in this plant in which the genome size is approximately three times the size of the human genome.
Asadolah Aslani Aslamarz, Kourosh Vahdati, Majid Rahemi, Darab Hassani and Charles Leslie
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’).
Kourosh Vahdati, Naser Lotfi, Bahman Kholdebarin, Darab Hassani, Reza Amiri, Mohammad Reza Mozaffari and Charles Leslie
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).