Showing papers in "Plant Cell Tissue and Organ Culture in 1991"
TL;DR: Shoot multiplication was achieved in vitro from shoot tips of aseptically germinated seedlings of Withania somnifera L. with direct multiple shoot initiation from germinating seeds in the presence of BA alone.
Abstract: Shoot multiplication was achieved in vitro from shoot tips of aseptically germinated seedlings of Withania somnifera L. using low concentrations of 6-benzyladenine (BA), viz. 2.2, 4.4 and 8.9 μM. Maximum number of shoots were obtained when 2.3 μM 2,4-dichlorophenoxyacetic acid (2,4-D) or 2.5 μM indolebutyric acid (IBA) was added to medium containing 4.4 μM BA during initiation of shoot multiplication, but not when added later. Direct multiple shoot initiation was also obtained from germinating seeds in the presence of BA alone. Rooting was successful in excised shoots grown on growth regulator-free MS medium. Rooted shoots were successfully established in soil in a greenhouse.
145 citations
TL;DR: An efficient adventitious shoot regeneration system was developed for pear, using leaves from in vitro proliferating shoots, and ammonium and total nitrogen proved to play an essential role: intermediate NH4+ concentrations were suitable for regeneration.
Abstract: An efficient adventitious shoot regeneration system was developed for pear (Pyrus communis L.), using leaves from in vitro proliferating shoots. Under optimal conditions, bud regeneration frequencies of ‘Comice’, ‘Passe-Crassane’, ‘Williams’ and ‘Conference’ ranged from 60% to 97%, with the mean number of shoots per regenerating leaf ranging from 3.2 to 6.6. Despite the great variability in responses of the different cultivars, in general an initial dark exposure of at least 20 days was required. Ammonium and total nitrogen proved to play an essential role: intermediate NH4
+ concentrations were suitable for regeneration. The balance between NH4
+ and NO3
- also influenced regeneration; optimal regeneration occured on media with a 1:3 NH4
+/NO3
- ratio. TDZ at 1 μM was less efficient than higher concentrations, whatever the NAA level. Finally, length and growth regulator composition of the two phases (induction and expression) influenced the regeneration rate of ‘Conference’.
144 citations
TL;DR: Chloroplasts of shootlets grown in a medium without sucrose contained numerous, large plastoglobuli and were lacking in starch granules, which might be used by the in vitro shootlets during the acclimation period.
Abstract: Shootlets of Rosa multiflora L. cv. Montse were cultured in vitro with four different levels of sucrose (0, 1, 3 and 5%). Chloroplasts of shootlets grown in a medium without sucrose contained numerous, large plastoglobuli and were lacking in starch granules. The size and number of starch granules increased with the level of sucrose in the culture medium. Starch content in leaves of shootlets grown with 5% sucrose was higher (ca 1, 3%) than those grown with 3% (ca 0, 45%) and 1% sucrose (ca 0, 27%). Starch might be used by the in vitro shootlets during the acclimation period.
129 citations
TL;DR: The content of internal cytokinins in the vitrified apple shoots was several times greater than in normal ones, which supports the hypothesis that excess of cytokinin, inducing rapid divisions of cells in meristems in the atmosphere with high humidity, is responsible for vitrification.
Abstract: Development of vitrification and apical necrosis was followed in Camellia sinensis, Gerbera jamesonii, Malus domestica and hybrid Populus tremula x P. alba shoots cultured in vitro on Murashige & Skoog (MS) medium with different concentrations of growth regulators. High humidity in the culture vessels and excess of BA in the medium were found to be the major factors influencing vitrification. Lack of exogenous cytokinin in the medium during successive subcultures induced apical necrosis in poor-rooting species (Malus domestica, Camellia sinensis). The level of internal phytohormones (ABA, IAA, IPA, 2iP, Z, ZR) was determined in the apple shoots by means of ELISA. The content of internal cytokinins in the vitrified apple shoots was several times greater than in normal ones, which supports the hypothesis that excess of cytokinins, inducing rapid divisions of cells in meristems in the atmosphere with high humidity, is responsible for vitrification. Apical necrosis of the plantlets that appeared after cultivation on cytokinin-free medium is the result of deficiency in endogenous hormones in apple shoots and this being confirmed by analysis of endogenous hormones in apple shoots.
126 citations
TL;DR: Although more shoots rooted in vitro than in vivo (77% vs. 46%), the latter was judged preferable since acclimatized plants were produced in less time and a major culture step was eliminated.
Abstract: Shoot apical meristems were used to establish regenerative axillary bud cultures of 9 muscadine grape cultivars. Meristems taken from 10 cm long shoots had less contamination (3%) and a higher survival rate (94%) than those from shorter or longer shoots. Of media tested, MS, 1/2 MS, and C2D resulted in equivalent shoot proliferation rates, whereas, WPM produced stunted shoots. When pooling results for 3 cultivars, 5, 10 and 20 μM BA and 5 μM TDZ produced the highest average number of shoots per cultured apex (3.4–3.8). However, shoots produced with TDZ were stunted and did not root well. For rooting of shoots directly in potting mix, a rooting powder pretreatment significantly increased the number of roots per shoot but did not affect percent rooting or root length. For rooting in vitro, 1 μM NAA significantly increased all parameters measured. Although more shoots rooted in vitro than in vivo (77% vs. 46%), the latter was judged preferable since acclimatized plants were produced in less time and a major culture step was eliminated. Significant differences among cultivars were noted for measured responses in all experiments.
100 citations
TL;DR: Callus was initiated from immature leaf and stem segments of rose and subcultured every four weeks on a basal medium of half-strength Murashige & Skoog (1962) salts plus 30 g l-1 sucrose (1/2 MS) and supplemented with 2.2 μM BA, 5.4 μM NAA and 2.0 μM 2,4-D.
Abstract: Callus was initiated from immature leaf and stem segments of rose (Rosa hybrida cv. Landora) and subcultured every four weeks on a basal medium of half-strength Murashige & Skoog (1962) salts plus 30 g l-1 sucrose (1/2 MS) and supplemented with 2.2 μM BA, 5.4 μM NAA and 2.2–9.0 μM 2,4-D. Embryogenic callus and subsequently somatic embryos were obtained from 8-week-old callus culture on 1/2 MS+2.2 μM BA+0.05 μM NAA+0.3 μM GA3+200−800 mg l-1 L-proline. Long-term cultures were established and maintained for up to 16 months by repeated subculture of embryogenic callus on L-proline deficient medium. About 12% of cotyledonary stage embryos taken from cultures cold-stored at 8±1°C for 4 days germinated on 1/2 MS+2.2 μM BA+0.3 μM GA3+24.7 μM adenine sulphate.
89 citations
TL;DR: Red beet cell lines exhibiting a range of cell colours were generated from secondary callus via specific induction methods, demonstrating that methylation plays a key role in the repression of genes encoding enzymes involved in betacyanin biosynthesis.
Abstract: Red beet cell lines exhibiting a range of cell colours were generated from secondary callus via specific induction methods. Phenotype colour ranged from white/green through yellow, orange and red to deep violet, representing all types of pigments found in red beet plant. Specific phenotypes could only be obtained through specific induction sequences and once established were stabilised by cultivation on a maintenance medium. The ratio of auxin (2,4-D) to cytokinin (6-BAP) was an important factor in the control of these processes. All coloured phenotypes were linked, but could be classified into two main groups, one yellow-red and the other orange-violet, according to their different cellular morphologies. A certain amount of instability still existed within each group. Modification of the growth regulator composition could be used to interchange specific combinations of coloured phenotypes, depending upon the initial state of cellular differentiation. Use of the DNA-methylation inhibitor 5-azacytidine demonstrated that methylation plays a key role in the repression of genes encoding enzymes involved in betacyanin biosynthesis. Furthermore, the poly(ADP-ribose) polymerase inhibitor 3-methoxybenzamide blocked the induction of the same gene set in a concentration dependent manner without affecting cell growth.
85 citations
TL;DR: Axillary bud development and adventitious bud formation was obtained with decapitated shoot explants of Aloe barbadensis Mill and more adventitious and axillary buds developed on nutrient media supplemented with IBA than with NAA.
Abstract: Axillary bud development and adventitious bud formation was obtained with decapitated shoot explants of Aloe barbadensis Mill. Maximal bud growth and rooting of shoots was obtained on a modified medium of Murashige and Skoog supplemented with 5 μM IBA. More adventitious and axillary buds developed on nutrient media supplemented with IBA than with NAA. Axillary buds but not adventitious buds developed with IAA in the medium. Morphogenesis was inhibited by 2,4-D. Kinetin, benzyladenine and thidiazuron were toxic to the explants and did not stimulate the development of axillary of adventitious buds. The optimal temperature for bud growth and development was 25°C. Axillary bud growth and the formation of adventitious buds was slowed down at 10°C and totally inhibited by 30°C. The optimal sucrose concentration was 3% with the inhibition of bud growth and development by higher sucrose levels.
79 citations
TL;DR: The Southern hybridization patterns indicate that in one cell line the T-DNA had undergone extensive rearrangements, and might indicate that the process of T- DNA transfer and integration might differ in the case of cereals.
Abstract: DNA can be transferred by Agrobacterium tumefaciens to wheat, albeit at very low frequencies. Transfer of agrobacterial DNA occurred in cultures where the embryos had been subjected to partial enzymatic digestion prior to cocultivation with the bacteria. It is unclear whether this is by the normal process mediated by the Ti virulence genes and the border repeats of the T-DNA. The Southern hybridization patterns indicate that in one cell line the T-DNA had undergone extensive rearrangements, and might indicate that the process of T-DNA transfer and integration might differ in the case of cereals. This could suggest the method of transfer and ultimately the expression of these genes in cereal cells may be different to that observed in other monocotyledonous and dicotyledonous species.
78 citations
TL;DR: No significant difference was observed in the number of embryos produced with 6% sucrose or 3% Sucrose plus an equimolar concentration of either mannitol, sorbitol, or myo-inositol in the maturation medium, suggesting that the effect of the carbohydrate on the m maturity was partly osmotic.
Abstract: Different carbohydrates were investigated for somatic embryo development of black spruce and red spruce. They were tested in a basal maturation medium consisting of Litvay's salts at half-strength containing 1 g l-1 glutamine, 1 g l-1 casein hydrolysate, 7.5 μM abscisic acid, and 0.9% Difco Bacto-agar. A comparison of different sucrose concentrations showed that 6% was optimal for embryo development. Among the nine carbohydrates tested, sucrose, fructose, glucose, maltose, and cellobiose supported embryo development while arabinose, mannitol, myo-inositol, and sorbitol did not. A comparison of sucrose, glucose, and fructose at three concentrations showed that the general pattern of response for both species followed concentration expressed as a percentage, independent of the molarity of carbohydrate in the medium. Interspecific differences were observed concerning carbohydrate requirements. For red spruce, 6% fructose was found best for embryo development, while no such preference was observed for black spruce. No significant difference was observed in the number of embryos produced with 6% sucrose or 3% sucrose plus an equimolar concentration of either mannitol, sorbitol, or myo-inositol in the maturation medium, suggesting that the effect of the carbohydrate on the maturation was partly osmotic.
76 citations
TL;DR: Explants excised from strawberry plantlets were cultured in vitro for 21 days on half-strength MS on basal liquid medium with 20 g l-1 sucrose and without sugar in the vessels capped with gas permeable microporous polypropylene film to study photoautotrophic and CO2 enriched conditions.
Abstract: Explants excised from strawberry (Fragaria x ananassa Duch.) plantlets were cultured in vitro for 21 days on half-strength MS (Murashige & Skoog 1962) basal liquid medium with 20gl 1 sucrose and without sugar in the vessels capped with gas permeable microporous polypropylene film. The experiments were conducted under CO 2 nonenriched (350-450/zmol mol 1 in the culture room) and CO 2 enriched (2,000/xmolmol -I during the photoperiod in the culture room) conditions with a PPF (photosynthetic photon flux) of 200/xmol m-: s -1. The CO 2 concentration in the vessels decreased to approximately 200/xmol mol -~ during the photoperiod on day 21 under CO 2 nonenriched conditions. The fresh and dry weight, net photosynthetic rate (NPR) per plantlet, NPR per g leaf fresh weight, NPR per g leaf dry weight, the number of unfolded leaves, and ion uptake of PO]-, NO3, Ca2+, Mg2+ and K + on day 21 were the greatest under photoautotrophic (no sugar in the medium) and CO 2 enriched conditions. The residual percent of PO]- was 3% on day 21 under photoautotrophic and CO 2 enriched conditions.
TL;DR: Histological studies revealed the origin of buds from the peripheral layers of the meristematic ring in green bean (Phaseolus vulgaris L.) plants regenerated from 3-day old seedling explants via organogenesis.
Abstract: Green bean (Phaseolus vulgaris L.) plants were regenerated from 3-day old seedling explants via organogenesis. The explants contained a cotyledon and a small portion (2–3 mm) of embryonic axis split in half. Explants were cultured on a defined medium containing glutamine as the sole nitrogen source. A ring of meristematic tissue was produced at the base of the axillary bud located at the cotyledonary node. The meristematic tissue was produced only if the axillary bud was present together with the cotyledon in the explant. Buds and shoots developed from the meristematic ring. Selected shoots produced roots when excised from the cluster of buds and transferred to root induction medium. Rooted shoots (plantlets) grew well and produced viable seeds when grown in the greenhouse. Histological studies revealed the origin of buds from the peripheral layers of the meristematic ring.
TL;DR: The dual role of sugar as carbon and osmotic source in shoot regeneration from leaf explants of egg-plant is discussed and sucrose or fructose at 44 mM was optimal in inducing shoot regeneration compared to sucrose.
Abstract: Uniform leaf pieces of egg-plant, Solanum melongena L., were cultured in Murashige and Skoog's medium containing 2 mg l-1 kinetin and varying sugar levels. Glucose or fructose at 44 mM was optimal in inducing shoot regeneration compared to sucrose. Sucrose at 11 and 22 mM induced more shoot organogenesis than at lower or higher levels. An additional 22 mM mannitol with 22 mM sucrose enhanced shoot regeneration significantly more than 22 mM sucrose alone. The dual role of sugar as carbon and osmotic source in shoot regeneration from leaf explants of egg-plant is discussed.
TL;DR: Cell cultures of grapes, Vitis vinifera L. cv Gamay Fréaux were grown under different conditions of external osmotic potential induced by an increase of sucrose concentration or by the addition of mannitol to the culture medium, resulting in a significant intracellular accumulation of anthocyanin especially peonidin 3-glucoside in the pigmented cells.
Abstract: Cell cultures of grapes, Vitis vinifera L. cv Gamay Freaux were grown under different conditions of external osmotic potential induced by an increase of sucrose concentration or by the addition of mannitol to the culture medium. Addition of 82 mM mannitol or increasing sucrose concentration to 132 mM had similar effects on repressing growth. Cyanidin 3-glucoside, peonidin 3-glucoside and peonidin 3-p-coumaroylglucoside are three main anthocyanins of Vitis cells. Increasing osmotic potential from −0.43 MPa to −0.8 MPa in the medium resulted in a significant intracellular accumulation of anthocyanin especially peonidin 3-glucoside in the pigmented cells. High osmotic potential appears to stimulate the methylation of anthocyanins. Osmotic potential is an important culture factor and may be useful in the controlling of anthocyanin production and composition.
TL;DR: Growth rates, secondary metabolite production and T-DNA structure of a number of hairy root lines were examined and found to be unchanged after cryopreservation, and the regeneration of proliferating roots from surviving root-tips was low.
Abstract: Crypopreservation methods were firstly developed for root-tips from hairy root cultures of Beta vulgaris, established after transformation by Agrobacterium rhizogenes. The effects of culture age, pre-growth, cryoprotection, freezing rate and post-freeze culture conditions were determined. The resulting freezing protocol was then used to cryopreserve transformed root cultures of Nicotiana rustica. Both species were viable after freezing (ca. 80%), according to fluorescein diacetate vital staining. However, on average the regeneration of proliferating roots from surviving root-tips was low (<20%). Growth rates, secondary metabolite production and T-DNA structure of a number of hairy root lines were examined and found to be unchanged after cryopreservation.
TL;DR: This review addresses the engineering consequences of the unique characteristics of plant cells on the scale-up of plant cell culture and predicts a renaissance in plant cellculture as a production strategy.
Abstract: The enormous versatility of plants has continued to provide the impetus for the development of plant tissue culture as a commercial production strategy for secondary metabolites. Unfortunately problems with slow growth rates and low products yields, which are generally non-growth associated and intracellular, have made plant cell culture-based processes, with a few exceptions, economically unrealistic. Recent developments in reactor design and control, elicitor technology, molecular biology, and consumer demand for natural products, are fuelling a renaissance in plant cell culture as a production strategy. In this review we address the engineering consequences of the unique characteristics of plant cells on the scale-up of plant cell culture.
TL;DR: During studies to optimize production of morphogenic callus from cultured leaf discs of sugarbeet, large differences were observed associated with the gelling agent employed, and water availability, as determined mainly by gel matric potential, was found to be the dominant factor.
Abstract: During studies to optimize production of morphogenic callus from cultured leaf discs of sugarbeet (Beta vulgaris L.) large differences were observed associated with the gelling agent employed. Water availability, as determined mainly by gel matric potential, was found to be the dominant factor. A simple method was devised to measure the relative matric potential of different gels. A precisely moistened filter-paper disc was placed on the gel surface, allowed to equilibrate, removed and weighed. The relative gain or loss of water from the paper disc was a measure of the matric potential of the gel and varied with both gel type and concentration. Leaf disc expansion and production of callus-derived embryos and shoots were shown to be directly proportional to gel matric potential. Water availability may also be affected by the ease with which liquid is expressed from gels in response to localized pressure caused by explant expansion and contortion. This property, called gel expressibility, was easily measured with a weight and capillary pipette and shown also to vary with gel type and concentration. Validity of the technique for measuring relative matric potential was verified physiologically by culturing leaf discs on filter-paper overlays to eliminate expressibility differences among gels. Additionally, comparison of leaf disc growth on uncovered gel surfaces versus filter-paper overlays demonstrated the contribution of liquid expression to overall water availability. Expression of liquid by explants on uncovered gel surfaces greatly enhanced the production of morphogenic callus.
TL;DR: Multiple vegetative shoots and flowers emerged from the nodes of explants derived from young seedlings of the bamboo Dendrocalamus hamiltonii when cultured on a Murashige & Skoog based medium supplemented with the cytokinin 6-benzyladenine.
Abstract: Multiple vegetative shoots and flowers emerged from the nodes of explants derived from young seedlings of the bamboo Dendrocalamus hamiltonii when cultured on a Murashige & Skoog based medium supplemented with the cytokinin 6-benzyladenine (BA) The largest number of shoots produced from intact cultured epicotyl tissue (ca 8 per explant) were obtained after 12 weeks on a medium containing 44 μM BA The first flowers were observed in vitro after only 13 weeks from initial germination of the seedlings The highest frequency of flowering (47% of nodal explants) was obtained when cultured explants were transferred from 222 μM BA to a growth regulator-free medium after 8 weeks
TL;DR: The growth retardant chlormequat stimulated microtuber formation by a recalcitrant cultivar of potato (Solanum tuberosum), but reduced microTuber fresh weight in a cultivar that tuberised readily in its absence.
Abstract: The growth retardant chlormequat stimulated microtuber formation by a recalcitrant cultivar of potato (Solanum tuberosum), but reduced microtuber fresh weight in a cultivar that tuberised readily in its absence. Inhibition of microtuber growth by high concentrations of chlormequat was confirmed using a different in vitro system where all cultivars tuberised in the absence of growth retardants. Alternative growth retardants were tested. Daminozide also had a detrimental effect on microtuber fresh weight, but ancymidol and paclobutrazol did not inhibit microtuber growth at the concentrations required for stimulation of tuberisation by recalcitrant cultivars. In addition, 10-5 M ancymidol and paclobutrazol inhibited premature sprouting of microtubers in vitro.
TL;DR: Desiccation tolerance of celery (Apium graveolens L.) somatic embryos was increased by supplementation of embryo-production medium with 1 μM abscisic acid (ABA) or 1 mM proline, with highest survival obtained with a combination of ABA and proline.
Abstract: Desiccation tolerance of celery (Apium graveolens L.) somatic embryos was increased by supplementation of embryo-production medium with 1 μM abscisic acid (ABA) or 1 mM proline, with highest survival obtained with a combination of 1 μM ABA and 1 mM proline. Addition of ABA and proline increased fatty acid accumulation by somatic embryos; the effect on fatty acid composition was inconsistent. Somatic embryos capable of germination differed from mature zygotic embryos by greater size, lower fatty acid level, and substantially lower proportion of oleic acid (18:1) as compared to linoleic acid (18:2).
TL;DR: Apical flower buds of Cymbidium goeringli Reichenbach fil were explanted on modified Murashige & Skoog medium supplemented with N6-benzyladenine and α-naphthaleneacetic acid and found to be optimal for initiating rhizome development and subsequent plantlet regeneration.
Abstract: Apical flower buds of Cymbidium goeringli Reichenbach fil. (ca 2 mm long) exeised from infloreseences (ca 5 cm long) were explanted on modified Murashige & Skoog medium (=MS medium) supplemented with N6-benzyladenine (BA) and α-naphthaleneacetic acid (NAA). Within 107 days of culture, swelling growth, chlorophyll synthesis, and subsequent rhizome differentiation were observed. MS medium containing 0.1 mg l-1 BA and 10 mg l-1 NAA was found to be optimal for initiating rhizome development and subsequent plantlet regeneration. Explants cultured on MS medium supplemented with 1 mg l-1 NAA alone formed a mass of rhizome branches. Multiple shoots of rhizome branches were induced from apical segments when rhizomes were transferred to MS medium containing 0.1 mg l-1 BA and 10 mg l-1 NAA.
TL;DR: Rose breeding is now open to the full range of in vitro genetic manipulation techniques involving protoplast technology, with high yields of protoplasts isolated from embryogenic suspension cultures of Rosa persica x xanthina and Rosa wichuraiana.
Abstract: High yields of protoplasts were isolated from embryogenic suspension cultures of Rosa persica x xanthina and Rosa wichuraiana using an enzyme mixture comprising 20 g l-1 cellulase ‘Onozuka’ R10, 1 g l-1 Pectolyase Y-23 and 10 g l-1 hemicellulase. Agarose-immobilized protoplasts gave the most consistent growth at a plating density of 5×104 protoplasts ml-1 on the basic medium of Kao & Michayluk (KM8p) containing 2 mg l-1 naphthaleneacetic acid and 1 mg l-1 benzylaminopurine. At 25°C in the dark, 0.004% of R. persica x xanthina protoplasts developed into colonies. Using similar culture conditions, but with a plating density of 9×104 protoplasts ml-1, 0.017% of R. wichuraiana protoplasts developed into colonies. On transfer of R. persica x xanthina colonies to Schenk & Hildebrandt's medium containing 3 mg l-1 2,4-dichlorophenoxyacetic acid, globular and later stage embryos were formed. Approximately 30% of these embryos developed into plantlets on transfer to basal Schenk & Hildebrandt's medium. Further development of the plantlets took place on cellulose plugs (Sorbarods) soaked in Murashige & Skoog's medium containing 0.05 mg l-1 naphthaleneacetic acid, 0.05 mg l-1 indole-3-butyric acid and 0.1 mg l-1 benzylaminopurine. Rose breeding is now open to the full range of in vitro genetic manipulation techniques involving protoplast technology.
TL;DR: The rate of corm growth on shoots cultured in a liquid medium supplemented with paclobutrazol and a saturating sucrose concentration, was a function of the length of the shoot's leaf blades, and was similar in light and in dark.
Abstract: Conditions were defined for precocious differentiation and improved growth of corms at the base of gladiolus shoots. Shoots were derived from explants cultured on agar solidified media, and corm regeneration was obtained in subsequent liquid shake cultures. Benzyladenine (BA), at 10-7 M, was found to have a stimulating effect mainly when provided to the shoots prior to manifestation of corm growth. Paclobutrazol and sucrose promoted corm formation when supplemented to the liquid media. Paclobutrazol, at 10 mg l-1, shifted assimilate allocation towards the growing corm. A differential promotion of corm development by sucrose was not observed, and the concentration of sucrose at which the sugar demand for maximal shoot and corm growth is satisfied (60 g l-1) was unaltered by the presence of paclobutrazol. The rate of corm growth on shoots cultured in a liquid medium supplemented with paclobutrazol and a saturating sucrose concentration, was a function of the length of the shoot's leaf blades, and was similar in light and in dark.
TL;DR: An Asiatic orchid was clonally propagated using seed-derived rhizomes as explants and auxin stimulated rhizome growth by increasing branching and fresh weight of the explant, with 1-naphthaleneacetic acid (NAA) being the most effective auxin.
Abstract: An Asiatic orchid, Cymbidium forrestii, was clonally propagated using seed-derived rhizomes as explants. The rhizomes were cultured and proliferated on Murashige and Skoog medium supplemented with various growth substances. Auxins stimulated rhizome growth by increasing branching and fresh weight of the explant, with 1-naphthaleneacetic acid (NAA) being the most effective auxin. All auxins tested suppressed normal shoot formation. The apical meristem of the rhizome reacted to exogenously applied auxin by reducing the cytoplasmic zone of the apical meristem and causing meristem derivatives to rapidly differentiate into vacuolated parenchyma cells. Leaf formation and development was retarded in the presence of auxin. Cytokinins generally reduced rhizome growth and the number of branches, but benzyladenine (BA) can induce shoot formation in vitro. BA induced the cytoplasmic zone of the apical meristem to enlarge and enhanced leaf development. A 5% (w/v) sucrose concentration was most effective in shoot induction when combined with 5 mg1-1 BA. Activated charcoal promoted rhizome growth; however, shoot formation was inhibited.
TL;DR: In calli from distal and proximal cotyledon segments and from radicles of mature wild olive zygotic embryos, rhizogenesis prevailed during the first and somatic embryogenesis during the second 30 day subculture period.
Abstract: In calli from distal and proximal cotyledon segments and from radicles of mature wild olive zygotic embryos, rhizogenesis prevailed during the first and somatic embryogenesis during the second 30 day subculture period. Rhizogenesis was enhanced by low IBA (0.5 and 2.5 μM) and 5.0 μM 2iP. By reducing by half the media salt concentrations and by inserting a 21 day dark period rhizogenesis was also enhanced. Somatic embryogenesis was inhibited by both 2iP and IBA at concentrations higher than 5.0 μM, but was not influenced by lower salt concentrations or by inserting the dark period.
TL;DR: A germinating-seed assay was developed to determine the susceptibility of dry bean (Phaseolus vulgaris L.) to infection by Agrobacterium tumefaciens and callus and root tissues, respectively capable of growing in the presence of kanamycin were obtained.
Abstract: A germinating-seed assay was developed to determine the susceptibility of dry bean (Phaseolus vulgaris L.) to infection by Agrobacterium tumefaciens. Seedlings infected one to three days after germination were more susceptible to A. tumefaciens infection than seedlings germinated for five to seven days and the galls that formed on the one to three day seedlings were significantly larger. Nineteen genotypes of dry bean were screened with this assay and all were equally susceptible to nopaline, octopine and agropine biotypes of A. tumefaciens. In addition, cotyledonary nodes and hypocotyls of P. vulgaris were inoculated with disarmed strain A. tumefaciens strain C58Z707 and the avirulent A. rhizogenes strain A4RS (pRiB278b), respectively. Both strains contain the binary plasmid pGA482 which has the neomycin phosphotransferase II (NPT II) gene nested between T-DNA borders. From these infected tissues, callus and root tissues, respectively capable of growing in the presence of kanamycin were obtained. These tissues displayed NPT II activity and integrated copies of the NPT II gene were detected from putative transformed root cultures by genomic blot hybridization.
TL;DR: An apparatus was developed that featured programmable application of liquid medium to plant cultures for micropropagation that maintained sterility was a key constraint in the development of the plant tissue culture apparatus.
Abstract: An apparatus was developed that featured programmable application of liquid medium to plant cultures for micropropagation. Computer control capabilities included liquid medium introduction and medium depth within four culture vessels, medium application and removal on an assigned schedule, schedule adjustment during a culture period and medium replacement. The medium level was controlled using an accurate custom level-sensing technique consisting of thermistors and float switches. Seven-liter polycarbonate containers were modified and used as the culture vessels. Maintaining sterility was a key constraint in the development of the plant tissue culture apparatus.
TL;DR: Uptake of BA was linear throughout this culture period in Musa but slowed considerably in Rhododendron shoots after day 10, and Benzyladenine was present in significant levels only in the pseudostem of Musa.
Abstract: Shoots of Musa and Rhododendron were cultured in vitro on a medium containing 05 mg l-1 BA Shoots growing in the presence of [14C]BA were harvested at intervals during the culture period Uptake of BA was linear throughout this culture period in Musa but slowed considerably in Rhododendron shoots after day 10 Rhododendron shoots absorbed 40% of the BA present in the medium and Musa shoots absorbed 52% In each species the principal metabolite formed was [9G]BA Benzyladenine was present in significant levels only in the pseudostem of Musa
TL;DR: Only 8 of 22 raspberry cultivars were capable of regeneration from leaf explants derived from greenhouse-grown plants, and TDZ was more effective than N-(phenylmethyl)-1H-purin-6-amine (BA) at promoting regeneration in combinations tested with IBA.
Abstract: Adventitious shoot regeneration was observed using leaf-petiole explants from shoot-proliferating cultures of ‘Comet’ red raspberry (Rubus idaeus L.). A maximum regeneration rate of 70% (3.7 shoots/explant) was obtained using 4.5–9.1 μM (1–2 mg l−1) N-phenyl-N′-1,2,3-thiadiazol-5-ylurea (thidiazuron or TDZ) with 2.5–4.9 μM (0.5–1 mg l−1) 1H-indole-3-butanoic acid (IBA) or 2.3 μM (0.5 mg l−1) TDZ with 4.9 μM (1 mg l−1) IBA in modified Murashige-Skoog medium. TDZ was more effective than N-(phenylmethyl)-1H-purin-6-amine (BA) at promoting regeneration in combinations tested with IBA (maximum 50% regeneration rate; 1.8 shoots/explant). Variation in the agar concentration or incubation temperature, orientation or scoring of the leaf-petiole explants and use of separate leaf or petiole explants had no effect on shoot regeneration. Incubation in the dark for 1, 2 or 3 weeks prior to growth in the light did not influence the percent regeneration rate but depressed the number of adventitious shoots. Explant source, from micropropagated shoots or greenhouse-grown plants, had an effect on shoot regeneration that was genotype dependent. Only 8 of 22 (36%) raspberry cultivars were capable of regeneration from leaf explants derived from greenhouse-grown plants.
TL;DR: Genotypic differences were found both in callus induction and somatic embryo formation, with I. pseudacorus responding better than I. versicolor or I. setosa in regenerated plants and two of the I. Pseudacorus regenerants were tetraploid.
Abstract: A method for plant regeneration of Iris via somatic embryogenesis is described. Root and leaf pieces from in vitro-grown plants of several genotypes of rhizomatous Iris sp. were cultured in vitro. Callus induction occurred only on root cultures incubated under low light intensity (35 μmol m-2 s-1) on two induction media containing 2,4-D (4.5 or 22.5 μM), NAA (5.4 μM) and kinetin (0.5 μM). Somatic embryos developed after transfer of callus onto four regeneration media containing 9 or 22 μM BA, or 5 μM kinetin and 2 μM TIBA or 9 μM BA and 4 μM TIBA. Plantlets could be obtained from these somatic embryos. Genotypic differences were found both in callus induction and somatic embryo formation, with I. pseudacorus responding better than I. versicolor or I. setosa. Cytological analysis performed on root tips of 80 regenerated plants revealed that two of the I. pseudacorus regenerants were tetraploid.