Hypericum perforatum L. (St. John's Wort) has an extensive history as an important medicinal herb used for the treatment of neurological and depressive disorders (Linde et al., 1996). The objective of this study was to establish an in vitro tissue culture protocol for St. John's Wort. Nodal segments, axillary buds, and leaf disc explants produced multiple shoots and callus on Murashige and Skoog minimal organics medium supplemented with combinations of indoleacetic acid (IAA; 0.57, 2.85, 5.71 μm) and benzylaminopurine (BA; 2.22, 4.44, 8.88 μm). Shoot production occurred on all combinations of IAA/BA tested and was significantly less in treatments without hormones. Callus production was higher on treatments containing 2.85 μm IAA + 4.44 μm BA, or 5.71 μm IAA + 8.88 μm BA. Shoots transferred to hormone-free medium at 8 weeks formed roots by 12 weeks. A micropropagation protocol was established for St. John's Wort using mature plants as the explant source.
Dormancy level is an important factor in rooting stem cuttings of conifers. Eldarica pine, a Mediterranean species, is a multiple flushing pine that does not appear to express endodormancy in southern New Mexico. Photoperiod manipulations can alter the dormancy level of some conifer species; however, effects on eldarica pine are unknown. Half-sib stock plants were randomly assigned to one of three photoperiods: natural daylength (>12 hours, control), long-term (7 months) exposure to 9-hour daylength (LTSD), and 2-week exposure to 9-hour daylength (STSD). Of the cuttings from LTSD stock plants, 78% rooted; however, only 67% of the cuttings from the other two treatments rooted. Differences in rooting also were related to shoot type of the cuttings. Cuttings from expanded short shoots without a bud rooted more frequently than cuttings from branch shoots with or without a bud present. Applications of these results are discussed.
Grand crinum Lily is an ornamental plant which is considered to be one of the architect's favorite accent. plants and one that bloom most of the year. A protocol for plant propagation from inflorescence is described. Explants were excised from inflorescence at the primordial stage. and cultured on Murashige and Skoog medium (MS: alone or supplemented with benzyladenine 4.4 or 13.3 uM benzyladenine (BA) and 0.5 uM naphthaleneacetic acid (NAA) and incubated for four weeks. Explants cultured on BA-containing media produced white flower-like structures on the receptacle which produced multiple shoots after additional four weeks on a fresh medium containing 4.4 uM BA and 0.05 uM NAA. Shoots were transferred to a fresh medium for further growth during which the basal stem reached 3 to 5 mm diameter. At this stage shoots, with or without roots, were transferred to soil, without acclimatization, and normal plants were established in soil.
Hypocotyl explants of three cultivars of melon (Cucumis melo L.) (cvs. Revigal, Topmark and Kirkagac), and a cucumber (C. sativus L. cv. Taoz) rapidly directly regenerated multiple shoots on Murashige and Skoog medium augmented with 4.4 μm benzyladenine. Regeneration from the hypocotyl resulted in nearly 100% diploid shoots, whereas regeneration from the cotyledons resulted in 40% to 70% polyploid regenerants. Regeneration from cotyledon explants of melon cv. Revigal required light, whereas regeneration from hypocotyl explants of melon cv. Revigal occurred in both light and darkness. Direct regeneration also occurred from the hypocotyl of cucumber cv. Taoz in both light and darkness, even though cotyledonary explants did not regenerate buds or shoots under the same conditions. This is the first report of regeneration from the Cucumis genus producing a fully diploid plant population.
Multiple shoots were obtained from shoot tips (2 to 3 mm) derived from mature plants (5 to 6 years old) of Citrus reticulata Blanco cv. Khasi mandarin and C. limon Burm.f. cv. Assam lemon when cultured on Murashige and Skoog (MS) medium, supplemented with (mg·liter-1) 1.0 BAP, 0.5 kinetin, and 0.5 NAA. Root induction was observed when 7-week-old single shoots (≈ 2 cm long) of both Citrus species were cultured on MS medium supplemented with (mg·liter-1) 0.25 BAP, 0.5 NAA, and 0.5 IBA. These plantlets were successfully established in the soil. Chemical names used: naphthalene acetic acid (NAA), indole 3-butyric acid (IBA), and benzylamino purine (BAP).
Regeneration in vitro from the embryonic axis in Phaseolus sp. has not been reported. Two embryo sizes, 0.3-0.4 mm and 0.6-0.7 mm long at 10-12 and 21 days after pollination, respectively, were excised from 4 P. vulgaris (P.v.) and 2 P. acutifolius (P.a.) genotypes. The embryonic leaves and radicale were removed, and 0.1-0.2 mm of the embryonic axis was cultured on Gamborg's B5 medium with 0, 5, 10 and 20μ MBA. The cultures were incubated in the dark at 25°C for 2 weeks followed by 1 week in continuous cool white light (25μ MS-1m2) before transferring to the second medium (0, 2μ MBA and 2μ MBA + 4μ MGA3). The tissues from the larger embryos initiated a single shoot without PGR in 30% of 1 P.v. explants and 30-60% in 2 P.a. The other 3 P.v. formed roots only. Multiple shoots were initiated in all P.v. (15-60%) and in 2 P.a. (60 and 70%) with 5 or 10μ MBA. The tissues from the smaller embryos had single shoots for all genotypes (30-60%) without PGR. Multiple shoots were initiated in 50-80% and 75-90% of the explants from P.v. and P.a., respectively, with 5 or 10μ MBA. Excess callus formed with 20μ MBA and regeneration decreased. After 3 weeks on the second medium, 6-8 shoot s/P. v. and up to 15-20 shoots/Pa. explants were observed.
Near-infrared (NIR) reflectance spectroscopy was used to determine the chemical composition of fruit and nut trees. Potted almond and bench-grafted Fuji/M26 trees were fertigated during the growing season with different N levels by modifying the Hoagland to create different levels of nitrogen and carbohydrates in plant tissues during dormancy. Dried, ground, and sieved shoot, shank, and root samples were uniformly packed into NIR cells and scanned with a Foss NIRSystem 6500 monochromator from 400 to 2500 nm. Statistical and multiple linear regression methods were used to derive a standard error of performance and the correlation between NIR reading and standard chemical composition analysis (anthrone, Kjedahl and Ninhydrin methods for carbohydrate, total N, and amino acid analysis, respectively) were determined. The multiple determination coefficients (R 2) of apple and almond tissues were 0.9949 and 0.9842 for total nitrogen, 0.9971 and 0.9802 for amino acid, and 0.8889 and 0.8687 for nonstructural carbohydrate, respectively.
Clonal propagation of pawpaw is currently limited to budding and grafting. A tissue-culture system to rapidly produce clonal material would be valuable for both production and preservation of germplasm. Forced scion wood, shoots from root cuttings, and seedlings were explant sources for ontologically mature, intermediate, and juvenile ages, respectively. Preliminary data indicated that nodal explants had more rapid adventitious shoot formation than shoot tip explants. Disinfestation protocols were developed for each explant source. Nodal explants were cultured on MS medium supplemented with 10 μM BA and 0.1 μM TDZ. Within 3 weeks, 60% of the seedling explants had expanded axillary buds, while no bud expansion was observed for explants of either the intermediate or mature sources. By 6 weeks, seedling axillary shoots had elongated and were suitable for subculture. By 8 weeks, multiple adventitious buds and shoots had formed on all seedling explants. At this same time, axillary shoots began to elongate on intermediate source explants, but mature source explants appeared to be recalcitrant. Explant exudation caused medium darkening, but, by reducing the transfer interval from 4 to 2 weeks, discoloration was minimized. Mature source explants were maintained in culture and after ≈7 months, axillary bud expansion occurred in a small percentage of these explants.
Several rose species (Rosa rugosa, R. wichuraiana, R. setigera, R. laevigata, R. banksiae, R. roxburghii, R. odorata and hybrids) were employed to establish the appropriate nutrient media for shoot multiplication and root initiation of cultured shoots and to describe a procedure for the successful transfer to soil of plants obtained in vitro. Cultured shoot tips and lateral buds from different genotypes proliferated multiple shoots on a basal medium (MS salt, vitamins, glycine, sucrose and agar) supplemented with 0mg/l to 6mg/l 6-benzylamino purine (BA) and 0mg/l to 0.5 mg/l naphthalene acetic acid (NAA). Most rose species cultured in a modified MS medium supplemented with 2mg/l BA showed good growth and shoot proliferation. The buds nearest the apex exhibited the slowest rate of bud development. Root development was enhanced and shoot development inhibited by lowering the concentration of MS salts to quarter- and half-strength. With difficult-to-root species, rooting was improved by supplementing the media with auxin or giving them 3-7days of dark treatment.
Abstract
A protocol for in vitro propagation was developed with two dry bean (Phaseolus vulgaris L.) cultivars. Shoot cultures were initiated by placing seedling shoot tips (1.0 to 1.5 cm) on Murashige and Skoog (MS) medium in which the effects of kinetin and BA alone or in combination with IAA or NAA were examined with regard to shoot multiplication and root or basal callus formation. The combination of BA (3.0 mg liter–1) and NAA (0.1 mg·liter–1) was most effective in shoot multiplication. At high concentrations of BA or kinetin (>10 mg·liter–1), shoot production and internode elongation decreased markedly and rosette-like cultures with multiple buds developed. Shoots were rooted on basal MS medium. Ramets grew to maturity in the greenhouse or field and produced fertile flowers, pods, and seeds. Chemical names used: N-(2-furanylmethyl)-1H-purin-6-amine (kinetin); N-(phenylmethyl)-1H-purin-6-amine (BA); 1H-indole-3-acetic acid (IAA); 1-naphthaleneacetic acid (NAA).