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.
Mohammad Sadat Hosseini Grouh, Kourosh Vahdati, Mahmoud Lotfi, Darab Hassani and Nejat Pirvali Biranvand
Azadeh Behrooz, Kourosh Vahdati, Farhad Rejali, Mahmoud Lotfi, Saadat Sarikhani and Charles Leslie
Drought stress is one of the main constraints limiting worldwide crop production. Arbuscular mycorrhizae (AM) and plant growth-promoting bacteria (PGPB) such as Azotobacter chroococcum and Azospirillium lipofrum have been shown to alleviate drought stress effects. Therefore, the interaction effect of AM fungi [Glomus mosseae, G. etunicatum, and a mix of these (G. mix), and PGPB bacteria (Azotobacter chroococcum + Azospirillium lipofrum)] was investigated in 1-year-old walnut seedlings (cv. Chandler) under normal and drought stress conditions. Drought stress reduced growth (plant height, root length, number of leaves, and fresh weight) and leaf nutrient content (N, P, and Zn) significantly of walnut plants. In contrast, proline, total soluble sugar, starch peroxidase enzyme activity, and total phenolic content of walnut leaves increased under this stress. Application of fungi or bacteria, and especially their simultaneous use, alleviated the negative effects of drought stress on walnut seedlings. AM fungi and PGPB increased significantly the content of some metabolites, including total phenolic content, proline level, peroxidase activity, total soluble sugar, and starch content as well as peroxidase enzyme activity. This led to an increase in walnut plant growth under the drought stress condition. Among AM fungi, G. etunicatum was more effective in reducing drought stress symptoms than either G. mosseae or the G. mix of fungi. In conclusion, use of G. etunicatum, along with PGPB, can reduce negative effects of drought stress on walnut seedlings.