highly efficient plant regeneration system through multiple shoot differentiation from commercial cultivars of barley (Hordeum vulgare L.) using meristematic shoot segments excised from germinated mature embryos

• Published in: Plant cell reports 2004-08, Vol.23 (1-2), p.9-16
• Main Authors: Sharma, V.K, Hansch, R, Mendel, R.R, Schulze, J
• Format: Article
• Language: English
• Subjects: Barley; Biological and medical sciences; Biotechnology; Buds; Cultivars; culture media; Embryos; Eukaryotic cell cultures; Fertility; Fundamental and applied biological sciences. Psychology; Gene Expression Regulation, Plant - genetics; Genotype; Genotypes; Hordeum - embryology; Hordeum - genetics; Hordeum - physiology; Hordeum vulgare; in vitro regeneration; Meristem - drug effects; Meristem - genetics; Meristem - physiology; Methods. Procedures. Technologies; micropropagation; Miscellaneous; Phenylurea Compounds; Picloram; Plant cells and fungal cells; Plant Shoots - drug effects; Plant Shoots - genetics; Plant Shoots - physiology; Plants, Genetically Modified - cytology; Plants, Genetically Modified - embryology; Plants, Genetically Modified - growth & development; Regeneration - drug effects; Regeneration - genetics; Seedlings; Seeds - drug effects; Seeds - genetics; Seeds - physiology; shoot meristems; Shoots; somatic embryogenesis; Thiadiazoles
• ISSN: 0721-7714; 1432-203X
• DOI: 10.1007/s00299-004-0800-4

A fast and highly efficient short-term in vitro regeneration system was developed for barley (Hordeum vulgare L.) based on readily available explants. Clumps of multiple shoots and buds suitable for transformation were obtained 9-10 weeks after culture initiation from model and current commercial cultivars. Meristematic shoot segments (MSSs) excised from mature embryo-derived seedlings and subsequently cultured on MS-based medium containing 2 mg/l Picloram and 3 mg/l thidiazuron (TDZ) differentiated up to ten multiple shoots after 3-4 weeks with no or very little callus formation. Sectors of the already multiplied shoot clumps were further multiplied on proliferation-maintenance medium containing 2 mg/l Picloram and 2.5 mg/l TDZ. Biweekly subcultures resulted in a continuous process of multiplication of these highly differentiating green sectors without any loss of morphogenic potential. The differentiated small shoots and shoot buds gave rise to normal shoots on medium with 0.1 mg/l Picloram and 1 mg/l TDZ. After rooting on basal medium with 0.5 mg/l or 1 mg/l IBA the plants were transferred to soil and showed normal growth and fertility compared to the seed-grown plants. All of the genotypes tested formed multiple shoots. The percentage of relative MSS multiplication was 63-83%, and the average number of multiplied shoots per MSS ranged from 16 to 34 among the genotypes after 9-11 weeks.