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This study was aimed at developing an efficient protocol for regeneration of Pseudostellaria heterophylla plantlets and induction of polyploidy. Calli of P. heterophylla (Miq) from stems, leaves and buds as explants could not differentiate into plantlets. However, young embryo segments used as primary explants produced embryonic calli on MS medium containing 5.0 mg/L 2,4-D and 0.5 mg/L KT. After the embryonic calli with granular protuberances were transferred to MS medium containing 0.5 mg/L BA, they developed shoots and then rooted to form plantlets. Polyploidy was induced when embryonic calli were placed in liquid MS medium containing 0.5% colchicine for 4 days, followed by culturing in solid medium to induce differentiation. Polyploidy was identified by the number of chromosomes and the size of plantlet stomata. The tetraploid plantlets produced larger root tubers than the diploid plantlets.
Solanum dulcamara L. (bittersweet) is a medicinal plant that has been used to treat skin diseases, warts, tumors, felons, arthritis, rheumatism, bronchial congestion, heart ailments, ulcerative colitis, eye inflammations, jaundice and pneumonia. A reliable in vitro culture protocol for bittersweet was established. Explants (leaf and petiole segments) were cultured on Murashige and Skoog minimal organics (MSMO) medium with various plant growth regulator combinations. Leaf explants formed more shoots than petiole explants. Plant regeneration was observed through indirect organogenesis with both explants. Best shoot proliferation was obtained from leaf explants with 3 mg/l BA (benzyladenine) and 0.5 mg/l IAA (indole-3-acetic acid). Regenerated shoots were transferred to rooting media containing different levels of IAA (indole-3-acetic acid), IBA (indole-3-butyric acid), NAA (naphthalene acetic acid) or 2,4-D (2,4 dichlorophenoxyacetic acid). Most shoots developed roots on medium with 0.5 mg/l IBA. Rooted explants were transferred to vermiculate in Magenta containers for acclimatization and after 2 weeks, they were planted in plastic pots containing potting soil and maintained in the plant growth room.
We aimed to produce tissue cultures and plant regeneration from endangered Crocus species: C. scepusiensis, C. tommasinianus, C. vittatus (“Verni” series of the genus) and C. banaticus. For initiation of cultures we used a plant growth regulator (PGR) combination used for in vitro culture of saffron and its relatives: 10 mg L-1 α-naphthaleneacetic acid (NAA) and 1 mg L-1 6-benzyladenine (BA). Shoot tips of young seedlings (C. scepusiensis) and corms (for the rest of species) were used as explants. C. scepusiensis explants developed into organogenic calli. On media with decreased NAA and with or without increased BA concentration, calli produced stigma-like structures and/or shoots and whole plants. In the other species, callus initiation medium induced callus formation with abundant somatic embryos. In C. tommasinianus, embryos developed shoots when auxin content of medium was decreased. In C. banaticus, a decrease of auxin with or without an increase in cytokinin content led to shoot or whole plant regeneration, as in C. scepusiensis. In the case of C. vittatus and C. banaticus, initiation and/or maintenance of cultures on indole-3-butyric acid (IBA) and increased sucrose concentration stimulated whole plant regeneration and in vitro cormlet development. C. scepusiensis and the rest of cultures (organogenic vs. embryogenic) differed at the biochemical level: C. scepusiensis cultures had higher (yet still low) enzymatic antioxidant (catalase, peroxidase) activities. With respect to catalase isoenzyme patterns, C. banaticus was different from the rest of cultures, demonstrating its distinct taxonomical position. Besides germplasm preservation use of the present cultures, they have a potential biotechnological value.
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