The role of nodal roots in prostrate clonal herbs: ‘phalanx’ versus ‘guerrilla’

The influence of nodal rooting on branching was studied in three evolutionarily and morphologically diverse species of prostrate clonal herbs: Tradescantia fluminensis (a monocotyledonous extreme ‘phalanx’ species), Calystegia silvatica (a dicotyledonous extreme ‘guerrilla’ species) and Trifolium repens (a dicotyledonous intermediate species). In all three, branch development from axillary buds is regulated by a positive signal produced by roots together with inhibitory influences from both pre-existing branches and shoot apical buds (apical dominance). Responses to nodal roots are cumulative and increased root activity leads to more vigorous bud outgrowth. In the absence of nodal roots, a single basal root system is unable to maintain continued extension growth of the shoot. We suggest that as individual nodal roots and stem internodes are both short-lived in these nodally-rooting clonal species, the plants’ investment in them is minimal. Thus, in contrast to perennial species lacking nodal roots, individual root systems in prostrate clonal herbs are small and stems have little secondary thickening and development of long-distance transport tissues. Hence the decline in extension growth of the shoot in the absence of nodal roots could be linked to the weak development of long-distance transport tissues in their relatively thin horizontal stems and to resource sharing between primary stems and lateral branches (as suggested by the greater retardation of primary stem growth in the more profusely branched ‘phalanx’ species (Trifolium and Tradescantia) than in the weakly branched ‘guerrilla’ species, Calystegia). These findings are consistent with the view that the long-term persistence of genotypes of nodally-rooting prostrate species is dependent upon them encountering the moist conditions required to facilitate the continual development of new young nodal root systems.     

Developmental and environmental regulation of anthocyanin pigmentation in wheat tissues transformed with anthocyanin regulatory genes

Summary Cell autonomous anthocyanin pigmentation, produced by the anthocyanin regulatory genes B and C1 controlled by the constitutive CaMV35s promoter (pBC1-7), was used to optimize biolistic gene delivery into embryogenic wheat (Triticum aestivum L. cv ‘Chris’) scutellum cultures. Intensely pigmented callus cells were observed 24 h postbombardment but these cells did not continue to divide and were developmentally terminal. A population of nonexpressing cells generated transgenic sectors which showed light-dependent anthocyanin pigmentation. Anthocyanin pigmentation was suppressed in regenerating shoot cultures but reverted to light-dependent production in the pericarp of developing seeds. Similarly, following microtargeted gene delivery into apical meristems, anthocyanin production was developmentally suppressed in leaf base meristems but prominent anthocyanin sectors developed in mature tissues beyond this region and persisted throughout leaf growth. In three developmental situations, callus proliferation, plant regeneration, and leaf growth, perpetuation of cells with anthocyanin regulator genes under the control of constitutive promoters was dependent on a higher level of regulation to suppress pigmentation at developmentally sensitive stages of meristematic activity. These findings provide additional evidence that the anthocyanin regulatory genes may be responsive to a variety of developmental and environmental stimuli. Present address: Genetics & Plant Breeding Department, G. B. Pant University of Agriculture Technology, Pantnagar, U.P., India, 263145.     

In vitro propagation of two species of commelinaceae from bud cultures and repeated subcultures on a growth regulator-free medium

Summary In our wide-ranging research on in vitro propagation of some monocotyledonous plants, two Commelinaceae species were studied: Tradescantia fluminensis var. foliis variegatis and Tradescantia pallida. Initial cultures were established successfully using nodal and apical meristems that produced single shoots, many roots, and no callus, by utilizing growth regulator-free MS medium. Addition of growth regulators did not increase the activity of explants that produced single or multiple shoots, atypical roots, and no callus. Consecutive cultures were possible using the apical and nodal meristems of the previous generation. The behavior of the different generations in culture did not change and was similar to the initial cultures. Their growth capacity was maintained over many months, also on a growth regulator-free medium. In both species, the chromosome number in the root tips of the mother plant and all morphologically stable in vitro plantlets confirmed a constant ploidy level, in T. fluminensis 2n=72, and in T. pallida 2n=24.     

Improved shoot organogenesis from hypocotyl segments of lingonberry (<Emphasis Type="Italic">Vaccinium vitis-idaea</Emphasis> L.)

Summary An improved protocol for shoot regeneration from hypocotyl segments of seedlings from open-pollinated seeds of lingonberry (Vaccinium vitis-idaea L.) cultivars, ‘Ida’, ‘Splendor’, and ‘Erntesegen’, and a native clone from Newfoundland was developed. The effect of thidiazuron (TDZ) on adventitious bud and shoot formation from apical, central, and basal segments of the hypocotyl was tested. Highly regenerative callus was obtained from hypocotyl segments on modified Murashige and Skoog (MMS) medium containing 5–10 μM TDZ. A maximum of 10 buds and 12 shoots per apical segment for seedlings of cultivar ‘Ida’ regenerated on MMS containing 10 μM TDZ. Callus and bud regeneration frequency, callus growth, and number of buds and shoots per regenerating explant depended not only on the specific segment of the hypocotyl, but also on parental genotype. Inhibition of shoot elongation by TDZ was overcome by transferring shoot cultures to a shoot proliferation medium containing 1–2 μM zeatin. The optimal concentration of sucrose for shoot elongation was 20 gl⁻¹. Shoots were rooted ex vitro on a 2 peat: 1 perlite (v/v) medium after dipping in 0.8% indole-3-butyric acid, and rooted plants acclimatized readily under greenhouse conditions.     

The initiation of callus and regeneration from callus culture ofTulipa gesneriana

Cold treatment of seeds, obtained from crosses between cultivars ofT. gesneriana L., affects the developmental stage of embryos, which in turn influences the frequency of callus induction and the development of different callus types. Cold-treated, mature embryos and basal segments ofin vitro-derived bulblets, were suitable explants for the initiation of regenerative callus on medium with 2,4-dichlorophenoxyacetic acid. The bulblets were initiated on flower-stalk segments from cold-stored bulbs ofT. gesneriana ‘Christmas Marvel.’ Histological analyses of regenerative callus revealed the regeneration of bulb-like structures. The influences of culture medium, culture conditions, growth regulators and acetylsalicylic acid, an inhibitor of ethylene, on the initiation and establishment of regenerative callus cultures are discussed.     

The establishment of cell suspension cultures ofGladiolus that regenerate plants

Summary Inflorencence stalks from greenhouse-grownGladiolus plants of the cultivars ‘Blue Isle’ and ‘Hunting Song’ cultured on a Murashige and Skoog basal salts medium supplemented with 53.6 μM 1-napthaleneacetic acid formed a compact, not friable type of callus that regenerated plantlets. Cormel slices and intact plantlets of three cultivars (‘Peter Pears’, ‘Rosa Supreme’, ‘Jenny Lee’) propagated through tissue culture formed a friable type of callus when cultured on Murashige and Skoog basal salts medium supplemented with 2,4-dichlorophenoxyacetic acid. This friable callus readily formed a cell suspension when the callus was placed in a liquid medium. Plants were regenerated from two-month-old suspension cell cultures of the commercial cultivar ‘Peter Pears’ after the suspension cells had been cultured on solid medium.     

The influence of plant growth regulators and storage on root induction and growth in <Emphasis Type="Italic">Pelargonium zonale</Emphasis> cuttings

The effects of post harvest application of ethylene, abscisic acid (ABA), indole-3-butyric acid (IBA) treatments or dark storage on root induction and continued growth of regenerated roots in Pelargonium cuttings were investigated using hydroponics in the greenhouse. Ethylene markedly increased rooting percentage in ‘Greco’ and ‘Surfing’, reduced the number of roots per cutting in ‘Surfing’ and had no effect on the total root lengths in the two cultivars. Ethylene treatment reduced fresh root mass in ‘Surfing’, increased dry root mass and reduced root water content in both cultivars. ABA (50 μM) enhanced rooting percentage in ‘Greco’, reduced the number of roots per cutting, reduced total root lengths and fresh root mass in both cultivars. ABA increased dry root mass and reduced root water content in ‘Surfing’ but this effect was not apparent in ‘Greco’. Storing cuttings in the dark for 4 days had no effect on rooting percentage and number of roots per cutting in ‘Greco’ and ‘Surfing’. However, dark storage reduced total root lengths in ‘Surfing’ and reduced fresh root mass in ‘Greco’. Dark storage had no effect on dry root mass and water content in both cultivars. Applying 4 μl l⁻¹ IBA in the rooting solution induced maximum (100%) root induction in ‘Surfing’. However, IBA reduced the number of roots per cutting in ‘Greco’, reduced total root lengths and fresh root mass in the two cultivars. IBA treatment profoundly increased and reduced dry root mass and root water content, respectively, in ‘Greco’ and ‘Surfing’. The enhanced root induction observed after IBA and ABA applications could be ascribed to their influence on ethylene biosynthesis, since ethylene treatment increased rooting percentage in both cultivars. However, high ABA (100 μM) and IBA (12 μl l⁻¹) levels or dark storage reduced the ability of induced roots to continue growth. We attribute our results to plant stress-response mechanism and ethylene appears to play an important role in the process of root initiation and root growth in Pelargonium cuttings.     

ABA and proline accumulation in leaves and roots of wild (<Emphasis Type="Italic">Hordeum spontaneum</Emphasis>) and cultivated (<Emphasis Type="Italic">Hordeum vulgare</Emphasis> ‘Maresi’) barley genotypes under water deficit conditions

The purpose of the study was to examine water stress-induced changes in the ABA and proline contents in roots and leaves of a potentially more resistant wild accession of Hordeum spontaneum and the modern cultivar Maresi (Hordeum vulgare). Leaves of H. spontaneum had higher contents of constitutive ABA and proline in comparison to those of ‘Maresi’. A moderate water deficit resulted only in root dehydration, which was higher in ‘Maresi’. Increases of water deficit in roots coincided with an increase of ABA content in roots, followed by that in leaves. The level of proline increased only in leaves and only in the case of H. spontaneum. Under conditions of severe water stress, the root dehydration levels were similar in the both genotypes, whereas leaf dehydration was higher in ‘Maresi’. H. spontaneum, as compared to ‘Maresi’ showed an earlier increase of ABA content in the roots and accumulated more ABA in the leaves. Free proline levels in the roots increased in both genotypes but H. spontaneum exhibited a 2-fold higher proline accumulation than ‘Maresi’. In H. spontaneum the accumulation of proline in the leaves occurred noticeably earlier and to a higher extent than in ‘Maresi’. A possible connection of these modifications with water stress resistance of the investigated genotypes is discussed in this paper.     

Response of durum wheat cultivars to immature embryo culture,callus induction and in vitro salt stress

Response of twenty eight cultivars of durum wheat (Triticum turgidum var. durum) to immature embryo culture, callus production and in vitro salt tolerance was evaluated. For assessment of cultivars to salt tolerance, growing morphogenic calli were exposed to different concentrations of NaCl (0, 0.3, 0.6, 0.9, 1.2, 1.5, 1.8 and 2.1% w/v) added to the culture medium during two subsequent subcultures (4 weeks each). Comparison of cultivars for callus induction from immature embryo was based on callus induction frequency and fresh weight growth of callus (FWG). While, for salt tolerance, the relative fresh weight growth (RFWG) and necrosis percent of callus were used. There were significant differences among cultivars for potential of regeneration from immature embryo, and ‘Shahivandi’ a native durum wheat cultivar originating from western Iran was superior among the cultivars tested. The FWG distinguished cultivars more than callus induction frequency did for callus induction evaluation. Hence, a range of FWG from 1.23 to 14.65 g was observed in ‘Mexical-75’ and ‘Omrabi-5’ cultivars, respectively. Growing calli derived from cultivars ‘PI 40100’ and ‘Dipper-6’ showed superiority for tolerating salinity under in vitro conditions. This revised version was published online in June 2006 with corrections to the Cover Date.     

In vitro propagation of Gladiolus hybridus Hort.: Synergistic effect of heat shock and sucrose on morphogenesis

Three cultivars (cvs.) of Gladiolus hybridus Hort., namely ‘Her Majesty’, ‘Aldebaran’ and ‘Bright Eye’ were successfully micropropagated. The cultures were established using intact cormels or segments of cormels and inflorescence axes on Murashige and Skoog (1962; MS) medium. The response depended on media supplements; both callus formation or direct induction of shoot buds was observed. Shoot differentiation from callus could be obtained on MS medium containing 1.0 μM BA (6–benzyladenine) and 10.0 μM NAA (α-naphthalene acetic acid) in all three cultivars. The same could be achieved by giving a heat shock (HS; 50 °C, 1h) to callus cultures (in case of ‘Her Majesty’ and ‘Aldebaran’ only) maintained on the basal medium. In these two cultivars, high sucrose concentration (0.232, 0.290 or 0.348 M) also favoured growth and proliferation of shoot cultures on a plant growth regulator-free medium at 20 °C in comparison to the cultures kept at 25 °C. On the other hand, shoot cultures maintained on the basal medium at 25 °C containing normal (0.058 M, i.e., 2.0%, w/v) sucrose concentration responded similar to those maintained at 20 °C on a high sucrose medium; reduced response was observed on normal sucrose containing medium at 20 °C. Heat shock enhanced shoot proliferation in the cultures maintained on basal medium, but induced prolific rooting in shoot cultures, within 5 days of HS, on high sucrose (optimum 0.232 M) medium. While the number of roots increased at higher sucrose concentrations in the medium in case of cvs. ‘Her Majesty’ and ‘Aldebaran’, the same was found to be independent of sucrose concentration in cv. ‘Bright Eye’. Generally the rooted plants produced on high sucrose (0.232 M) medium in comparison to medium with normal sucrose concentration showed better survival (ca. 90% as against 40%) in the soil. This revised version was published online in June 2006 with corrections to the Cover Date.     

Embryogenic mango cultures selected for resistance toColletotrichum gloeosporioides culture filtrate show variation in random amplified polymorphic DNA (RAPD) markers

Summary Genomic DNA isolated from embryogenic cultures of two mango cultivars, ‘Hindi’ and ‘Carabao,’ that had been selected for resistance to the culture filtrate ofColletotrichum gloeosporioides, was analyzed using Randomly Amplified Polymorphic DNA (RAPD).In vitro selection caused changes in RAPD markers in the selected embryogenic cultures with respect to the unchallenged control cultures and the stock plants. The differences involved both the absence and the presence of additional RAPD markers in the resistant lines, although the former was most commonly observed. The absence of differences between the unchallenged control of either cultivar and DNA from the leaves of parent trees confirmed that the changes were not due to prolonged maintenance in liquid cultures.     

Rapid production of wheat cell suspension cultures directly from immature embryos

The aim of this study was to produce suspension cultures of winter wheat directly from immature embryos bypassing the callus stage, and to determine their capacity for growth and regeneration in comparison to suspension cultures produced from callus. The study was carried out using Polish winter wheat varieties: ‘Grana’ and ‘Rosa’. Immature embryos were isolated, homogenized and transferred directly to liquid medium supplemented with 2,4-D. Actively dividing cell cultures were obtained within 2 months after the cultures were started. Suspension cultures from callus of immature embryos was also produced. With both cultivars, faster growth was observed in the suspension cultures produced directly from embryos than in the suspensions produced from callus. Metabolic activity was higher in the suspension culture produced directly from embryos than in the suspension derived from callus only in ‘Grana’. The production of 1-amiocyclopropane-1-carboxylic acid (ACC), an ethylene precursor, was lower in the suspension cultures produced directly from embryos than in the suspensions produced from callus. Morphogenic capacity was significantly higher in aggregates derived directly from embryos than in aggregates derived from callus. With ‘Rosa’, about one third of the aggregates derived directly from embryos regenerated shoots. Production of ACC was lower in ‘Rosa’ cell culture that regenerated then in other cell cultures that did not. Photosystem II reactions were more efficient in dark green aggregates than in light green or pale green aggregates which were unable to regenerate. With the method presented, wheat cell suspension cultures with a regeneration potential can be produced in 2 or 3 months less time than with traditional methods.     

Excessive aluminium accumulation in the pea mutant E107 (brz)

E107 is a pleiotropic mutant of peaPisum sativum cv. ‘Sparkle’, characterized by forming few nodules and developing bronze necrotic spots on older leaves. The mutant accumulates Al and has symptoms typical of Al toxicity. The lateral roots of E107 are fewer (40%) and shorter (50%) than those of its parent. High concentrations of Al accumulate in E107 shoots (1000 mg kg^-1) and roots (3000 mg kg^-1), and three-week old E107 plants extrude 2.5 times more protons than ‘Sparkle’ plants of similar age. Al concentrations of the roots of the mutant and of its parent ‘Sparkle’ are similar for the first two weeks of growth. Thereafter they differ. In 2 week old plants Al continues to accumulate in excessive amounts in E107 primary and lateral roots whereas in ‘Sparkle’ roots, it reaches a plateau. In E107, Al is erratically distributed in the walls of root hairs and epidermal cells in both primary and lateral roots. Some of these cells have also Al in their nucleus.     

In vitro assay of native Iranian almond species (<Emphasis Type="Italic">Prunus</Emphasis> L. spp.) for drought tolerance

Eight native Iranian almond species from three sections, ‘Euamygdalus’ (Prunus communis; Prunus eleagnifolia and Prunus orientalis); ‘Lycioides’ (Prunus lycioides and Prunus reuteri) and ‘Spartioides’ (Prunus arabica, Prunus glauca and Prunus scoparia) were in vitro screened for drought tolerance using sorbitol and polyethylene glycol (PEG) as an osmoticum. Different levels of water stress were induced using five concentrations of either sorbitol or polyethylene glycol in Woody Plant Medium (WPM). Water potential of various media ranged from −0.80 to −2.05 MPa and water stress in culture medium adversely affected plantlet growth. Wild species from ‘Spartioides’ were less affected than ‘Lycioides’ and ‘Euamygdalus’. At the same level of water potential, sorbitol had lower adverse effects than PEG; the latter being severe. Prunus × sorbitol and Prunus × PEG interactions were significant. At 0.2 M sorbitol and 0.003 M PEG, ‘Spartioides’ produced significantly more roots with higher total root length and root volume, as well as root-dry weight than those of ‘Lycioides’ and ‘Euamygdalus.’ It is concluded that in vitro screening of native Iranian almond species under specific and limited water-stress conditions may provide a system for effectively differentiating the wild species of almond for their expected root mass production under field conditions.     

The development of an efficient cultivar-independent plant regeneration system from callus derived from both apical and non-apical root segments of garlic (<Emphasis Type="Italic">Allium sativum</Emphasis> L.)

Summary Callus induction and later plant regeneration were studied in four widely grown garlic (Allium sativum L.) cultivars from Europe. Root segments from in vitro plantlets were used as starting material. In addition to cultivar effects, the effects of auxin and cytokinin levels and the position of the segments on the root were studied. There were no statistically significant differences among cultivars for the number of root segments that induced callus in the two series of experiments. The average induction frequency was 34.7% in the first series of experiments. Callus induction on apical root segments was significantly higher compared to callus induction on non-apical root segments in the second series of experiments. Two months after callus induction, callus lines were transferred to a regeneration medium consisting of Murashige and Skoog basal medium supplemented with 30gl⁻¹ sucrose and 1 mgl⁻¹ (4.6μM) kinetin. Calluses derived from different experiments were quite uniform with respect to their regeneration potential. Also it was found that our regeneration system was cultivar-independent. The average shoot regeneration frequency was 17.9% in the first series of experiments. Highly significant differences were found in the frequency of shoot regeneration among different callus induction treatments. When the cytokinin 6-(γ,γ-dimethylallylamino)purine (0.1mgl⁻¹∶0.5 μM) was present during callus induction, shoot regeneration ranged from 30.10 to 47.60%. Shoot regeneration from callus induced on non-apical segments was higher, although not significant, compared to callus induction from apical root segments in the second series of experiments. All in all, an efficient callus induction and plant regeneration system was developed from both apical and non-apical segments taken along the entire length of the roots. This system has potential to be used for garlic transformation.     

Adventitious shoot regeneration from Sinningia speciosa leaf discs in vitro and stability of ploidy level in subcultures

Summary Leaf explants of Sinningia speciosa were cultured in vitro on Murashige and Skoog (MS) basal medium with various growth substances in order to regenerate shoots. On MS medium supplemented with indoleacetic acid (IAA) and kinetin, 80% of the explants produced green callus and 25 to 30 shoots with roots per explant. On MS supplemented with IAA and N⁶ benzyladenine (BA), 80% of the explants produced green callus and 40 to 50 shoots per explant but lacked roots. After 3–4 mo., these shoots were removed from the initial explants and transferred separately onto MS supplemented with indolebutyric acid for their elongation and successive rooting (3 mo.). Histological studies showed that the callus was associated with mesophyll cell layers, primarily with the spongy parenchyma. The shoots regenerated at the callus surface and were associated with newly differentiated vascular areas. Recurrent regenerations were obtained from leaf explants or apical meristems excised from shoots of the previous subcultures. These explants, as compared to initial cultures, had a high frequency of regeneration and also produced more shoots per explant. Chromosome numbers of root tip cells of the mother plant and of all in vitro-regenerated plants remained constant: 2n=26.     

Assessment of a bacteriocin-producingLactobacillus strain in the control of spoilage of a cereal-based African fermented food

As part of a program to develop starter cultures aiding in the spoilage control and sanitation of African fermented foods, a cereal-based food (‘ogi’ and its solid form ‘agidi’ or ‘eko’) was prepared using a bacteriocin-producingLactobacillus strain as the starter culture. The survival of an enterotoxigenicEscherichia coli strain was investigated in the naturally fermented food and in food fermented with the starter bacteriocin-producingLactobacillus strain. An inhibition ofE. coli was observed within 2 h of incubation in ‘ogi’ fermented with the bacteriocin producing strain. After 6 h, the viable count ofE. coli in locally fermented ‘ogi’ was log 6.41 (2.54×10⁶CFU/mL), whereas in ‘ogi’ fermented with the bacteriocin producer it was reduced to log 1.70 (0.5×10² CFU/mL). Comparison of the shelf life of ‘agidi’ prepared from the naturally fermented food with that fermented with the bacteriocin-producing starter culture showed that the latter had a better shelf life (kept for 11 d before spoilage occurred as compared with 7 d for the natural one). The results are discussed in terms of the potential of bacteriocin-producing cultures in the control and retardation of spoilage and food-forne infections in some African fermented foods.     

Long-term study of somatic embryogenesis from anthers and ovaries of 12 grapevine (<Emphasis Type="Italic">Vitis</Emphasis> sp.) genotypes

Summary Anthers and ovaries of six grapevine cultivars (three Vitis vinifera L., two V × Labruscana L. H. Bailey, and one complex hybrid) were extracted from flower buds over 2 yr and cultured on three media reported to promote somatic embryogenesis in Vitis tissues. The highest percent embryogenesis from the hybrid ‘Chancellor’ and V. vinifera ‘Chardonnay’, ‘Merlot’, and ‘Pinot Noir’ occurred on medium C [Nitsch and Nitsch, 1969, basal medium with 3.0% (w/v) sucrose, 0.01% (w/v) inositol. 0.3% (w/v) Phytagel, 2.5 μM 2.4-dichlorophenoxyacetic acid, 2.5μM β-naphthoxyacetic acid, 5.0μM N-(2-chloro-4-pyridyl)-N′-phenylurea, and 0.05% (w/v) glutamine]. Regardless of the media, the labrusca cultivars ‘Concord’ and ‘Niagara’ produced soft non-embryogenic callus that was sometimes mixed with well-developed somatic embryos. Nine vinifera genotypes were further tested for several different years on medium C. Embryogenic cultures suitable for transformation were obtained from all genotypes in more than 1 yr. The average percent embryogenesis from ovaries was 7-fold higher than from anthers. There was significant annual variation in percent embryogenesis, demonstrating the need for media comparisons to be replicated for more than one season. Suspension cultures suitable for use in genetic transformation were initiated from ‘Chardonnay’, ‘Merlot,’ and ‘Pinot Noir’ pro-embryogenic masses. ‘Chardonnay’ suspension cultures plated and grown under conditions developed for recovery of plants after biolistic transformation yielded approximately 500 non-transformed embryos per plate after 4 mo. of culture, with 68.6% of the embryos converting to plants. This is the first reported protocol for embryogenesis from ‘Concord,’ ‘Cabernet Franc,’ and ‘Pinot Noir’ grapevines.     

Effect of bialaphos and phosphinothricin on plant regeneration from long- and short-term callus cultures of Gladiolus

Summary Callus was initiated from in vitro-grown plants of Gladiolus cultivars ‘Jenny Lee’ and ‘Florida Flame.’ The age of callus used for regeneration of plants was either 9 mo. old or 8 yr old from ‘Jenny Lee,’ and 4 mo. old from ‘Florida Flame.’ Regeneration medium consisted of Murashige and Skoog’s basal salts medium supplemented with 2.0 mg/l (9.3 μM) kinetin. This medium was supplemented with various concentrations of either bialaphos (Meiji Seika, Tokyo, Japan) or phosphinothricin (Hoechst-Roussell, Frankfurt, Germany). Bialaphos was more effective than phosphinothricin at stimulating plant regeneration. Plants regenerated from 8-yr-old callus of ‘Jenny Lee’ only when the regeneration medium was supplemented with 0.10 mg/l bialaphos. A bialaphos concentration of 0.01 mg/l stimulated regeneration from 9-mo.-old callus of cultivar ‘Jenny Lee’ and 4-mo.-old callus of ‘Florida Flame.’     

Studies on anther cultures of tomato —Lycopersicon esculentum Mill

The anthers of three genotypes ofLycopersicon esculentum, viz. cv. HS-101, cv. HS-102 and an F₁ hybrid (Montfavet 63-4xHS-101) in different stages of development were cultured in various defined nutritive media. Only anthers containing microspores in the early uninucleate stage were found to respond with the culture medium in the formation of androgenic callus. The DGII medium with 2 mg l⁻¹ NAA and 1 mg 1⁻¹ kinetin was found to be best for callus induction but MS medium supplemented with 2 mg l⁻¹ 2,4-D and 0.1 mg 1⁻¹ BAP favoured proliferation and growth of the callus. The androgenic microspores followed the ‘B’ type pathway of androgenesis in the formation of callus. Induction of tracheids in the callus could be achieved by supplementing the basal medium with NAA and kinetin or 2,4-D and BAP. Initiation of vessel elements and cambium were favoured by addition of NAA and kinetin and that of the phloem in the presence of 2,4-D and BAP in the basal medium, suggesting that the hormonal requirements for production of different elements of the vascular system in androgenic callus are different. Although roots could be induced from the callus, shoot differentiation could not be achieved under cultural conditions.