Lilium Asiatic Research Articles

Plant regeneration via embryo-like structures: histological observations and genetic stability in regenerants of Lilium spp.

SUMMARYTransverse thin-cell layers (tTCLs) excised from bulblets of Lilium longiflorum X Oriental hybrid ‘Triumphator’ were cultured on 1.0X Murashige and Skoog (MS) medium containing 0.1 mg l−1 α-naphthaleneacetic acid (NAA) and 0.1 mg l−1 kinetin (KT) in the dark at 23° ± 2°C. White-yellow, friable, embryo-like structures formed after 6 weeks in culture. The embryo-like structures proliferated well when cultured on 1.0X MS medium containing 1.0 mg l−1 NAA and 0.2 mg l−1 thidiazuron (TDZ), and converted into whole plantlets on 0.5X MS medium without any added plant growth regulator. Histological studies identified the origin of the first cell divisions from single sub-epidermal cells, from which somatic embryos developed. Using this procedure, embryo-like structures (ELS) were obtained at frequencies of 85.5%, 76.8%, 68.4%, 28.5% and 25.6% for L. longiflorum, L. longiflorum X Oriental ‘Triumphator’, Lilium Oriental hybrid ‘Siberia’, Lilium Asiatic hybrid ‘Elite’, and L. davidii var. unicolor, respectively. Embryo-like structures could be proliferated and developed into plantlets, with five-to-ten plantlets per embryo-like structure in all five Lilium species and hybrids tested. No polymorphic bands were detected using inter-simple sequence repeat (ISSR) markers and no change in ploidy level was found by flow cytometry (FCM) among the regenerants of all five Lilium species and hybrids. These five Lilium species and hybrids represented a diverse genetic resource of the genus Lilium. This protocol may be widely applicable for somatic embryogenesis and has potential applications in micropropagation and genetic transformation in Lilium spp.

Vermicompost induced changes in growth and development of Lilium Asiatic hybrid var. Navona

Organic agriculture minimizes the negative effects of agricultural activities. The present study was carried out in the factorial experiment on the basis of Complete Randomized Design (CRD) with 4 treatments and 3 replications. The vermicompost used was prepared using bovine manure and the earthworm (Eisenia foetida). Applied vermicompost levels were V0 = 0%, V1 = 10%, V2 = 20%, V3 = 30%. The obtained results from the present research indicated that applied vermicompost especially, at 30% level had significantly improving effects on the accumulation of macro nutrients, Ca and K, and micronutrients, Zn and Fe in both stems and root tissues. Applied vermicompost resulted in better growth and development of vermicompost treated plants as they had higher number of leaves, leaf dry mass, fresh stem and dry weight, stem height and diameter, root number and length. Observed increased contents of gibberellic acid in root tissues resulted from the application of vermicompost. Vermicompost treatments at suitable levels of 20 and 30% had stimulating effects on the number of flowers and their diameters and reducing effects on time of flowering. The result of the present study showed that the use of vermicompost, particularly, with a 30% content had favorable results on the growth and development of the Lilium asiatic hybrid var. Navona. Key words: Biofertilizer, flowering, Gibberellins, humic acid, ornamental plant.

Cloning of SCA Gene Related to Pollen Tube Adhesion and Oriented Growth and Analysis of Gene Diversity in <i>Lilium spp.</i>

Stigma/style cysteine-rich adhesin genes ( SCA ) were cloned from ‘Tresor’ ( Lilium Asiatic Hybrids), ‘White heaven’ ( Lilium Longiflorum Hybrids) and ‘Caruso’ ( Lilium Oriental Hybrids) leaves by PCR approaches. In this research we obtained four gene copies in both of ‘Tresor’ and ‘Caruse’ genomes and three gene copies in ‘White heaven’ genome. The identities of the cloned SCA among the copies were in the ranges from 72.97% to 99.68%. There are two exons and one intron in all of cloned SCA sequence, which could be deduced immature protein with 113 or 115 amino acid residues (encoding mature protein with 91 amino acid residues) SCA protein contains an N-terminal signal peptide of typical LTP protein, eight conserved cysteine residues and two consensus pentapeptide motifs. The identities among the amino acid sequences were in range from 87.91% to 98.90% higher than that of the similarities of gene sequences. Alignment analysis of amino acid sequences of cloned SCAs showed there were some distinct differences of amino acid site existing in the tested three species of lily hybrids. The phylogenetic tree revealed the relationship of the cloned 11 amino acid sequences of SCA were all more close to LTP family of monocotyledons than that of dicotyledons, which were consistent with the conclusions of kinship in plant taxonomy.

Floral nectary structure, nectar production, and carbohydrate composition in theLiliumAsiatic hybrid ‘Trésor’

Floral nectary structure, nectar production, and carbohydrate composition were compared from petals (“inner tepals”) and sepals (“outer tepals”) of Lilium Asiatic hybrid ‘Trésor’ (Liliaceae). The six nectaries each occupied a narrow furrow bordered by two convergent ridges extending adaxially from the petal and sepal base. Each sepal nectary furrow was shorter and more concealed. In both nectary types, many vascular bundles comprising xylem and phloem supplied 5.5–8 layers of nectariferous parenchyma cells below the epidermis, which lacked stomata. Transmission electron microscopy of sepal nectaries demonstrated that parts of the outer epidermal wall adhered to an intact but uplifted cuticle in nectar-secreting flowers. Both apoplastic and symplastic routes were continuous from the vascular bundles to the nectary epidermis. Starch breakdown from amyloplasts throughout the nectary likely augmented nectar production. Nectar solute concentration from another Asiatic hybrid, ‘Orange Pixie’, was also significantly higher in petals. In ‘Trésor’, significantly more nectar was available from sepals, possibly reflecting reduced evaporation from multiple nectar droplets within the covered nectary furrow. However, for both hybrids, the same quantity of nectar sugar was produced by petals and sepals. Nectar composition from petals and sepals also was alike, in ‘Orange Pixie’ averaging 67/19/14 (= sucrose/fructose/glucose) and 59/25/17, respectively, and in ‘Trésor’ averaging 68/23/10 and 62/27/12, respectively.

Effect of Pulsing Treatments for Enhancing Shelf-Life of Cut Asiatic Lilium cv. Elite

Studies were conducted on cut Asiatic Lilium cv. Elite to assess the effectiveness of various floral preservatives as pulsing treatments for delaying senescence and prolonging vase life. Uniform spikes of lilium at bud colour break stage were brought to the laboratory in the morning and placed in 8 different pulsing solutions consisting of sucrose (Suc) 5%, aluminium sulphate (AS) 400 ppm, silver thio-sulphate (STS) 2.0 mM and citric acid (CA) 1000 ppm alone and in combination with sucrose. Distilled water without any chemical served as the control. Among individual treatments, STS 2.0 mM maintained better water relations and flower quality compared to others. STS also showed superiority over other treatments when combined with Suc 5% by providing largest flower size (16.74 cm) with maximum vase life (17.29 days) owing to most-favourable water relations parameters.

COMPARISON OF SILVER THIOSULFATE WITH 1-METHYLCYCLOPROPENE ON 19 CUT FLOWER TAXA

The effects of silver thiosulfate (STS; AVB) and 1-methylcyclopropene (1-MCP; Ethylbloc) were determined on 14 commonly-grown cut flower species, represented by one to three cultivars per species. Stems were unpacked, sorted, and placed in either deionized water (DI) and subjected to 1-MCP (740 nL L') or ambient air for 4 h or DI plus STS at either 0.1 mM (Alstroemeria) or 0.2 mM (all other species) for 4 h. After treatment, stems were removed, placed in polyethylene sleeves and stored either wet in DI water or dry in plastic-lined floral boxes at 5°C in the dark for 4 days. After storage bunches were placed in DI water under 12 h (76 to 100 μmol m -2 s -1 ) light per day. Flowers were monitored daily to determine the end of wholesale vase life, which was designated as the first day a change was noticed in the flower or inflorescence that would typically prevent it from being sold by a wholesaler or retailer. The consumer vase life was also recorded for each stem and was designated as the day a typical consumer would dispose of it. The 19 cut flower taxa could be organized into four groups based on effectiveness of STS and 1-MCP: (1) Both STS and 1-MCP increased vase life but STS was more effective: Dianthus caryophyllus (all three cultivars), Bouvardia, Lilium (Asiatic), and Lathyrus odorata. (2) Both STS and 1-MCP prevented the negative effects of dry storage: Freesia (both cultivars) and Chamelaucium (one cultivar). (3) STS increased vase life while 1-MCP did not: Alstroemeria, Delphinium, Matthiola, and Gypsophila. (4) STS and 1-MCP either had no effect or a negative effect: Consolida, Eustoma, Ranunculus, Antirrhinum, and Chamelaucium (one cultivar).

EFFECT OF CERTAIN PRE-TREATMENT SUBSTANCES ON VASE LIFE AND PHYSIOLOGICAL CHARACTER IN LILIUM SPP.

The effects of pre-treatment substances on vase life and physiological character of cut Lilium Oriental hybrid ‘Siberia’ and Lilium Asiatic hybrid ‘Dream Land’ were studied. When florets of ‘Siberia’ were treated with Promalin (GA4+7+BA), the vase life was increased as compared to other treatments. The amount of ethylene produced continuously decreased and respiration rate showed a climacteric type rise on the 5 th day. The sugar contents increased in petal, but decreased in leaf and ovary. When florets of ‘Dream Land’ were treated with GA4+7 and Promalin, the vase life increased. The amount of ethylene production and respiration rate showed peak points on the 1 st day. Also, the sugar contents showed the same results as with ‘Siberia’. In the case of ‘Siberia’, vase life was extended with Dan-solution 2 [(nano particle pure colloidal Ag + ion) 0.1% + Natural chitosan] treatment. The amount of ethylene produced continuously decreased and respiration rate showed a climacteric type rise on the 5 th day. But they were not significantly different between treatments. The sugar contents increased in petal, but decreased in leaf and ovary. When florets of ‘Dream Land’ were treated with Dan-solution 2, the vase life was also increased. The amount of ethylene produced and respiration rates showed peak points on the 1 st day. Also, the sugar contents showed the same results as with ‘Siberia’.

Sprouting rate, change of carbohydrate contents and related enzymes during cold treatment of lily bulblets regenerated in vitro

This study was performed to investigate the effect of low temperature storage on the dormancy of in vitro bulblets of Lilium Oriental hybrid ‘Casablanca’ and Lilium Asiatic hybrid ‘Mona’. All bulblets regenerated in vitro remained dormant, irrespective of cultivar, if they are not subjected to cold treatment. Physiological dormancy was released more rapidly and the highest percentage of sprouting occurred after a 9-week cold treatment at 4 °C, as compared with 25 and 10 °C. Exposure to low but non-freezing temperatures induced the net breakdown of starch and the accumulation of sucrose and glucose in lily bulblets regenerated in vitro (low temperature sweetening). Bulb sweetening was especially pronounced at 4 °C. Analysis of carbohydrate types revealed that sucrose was the dominant soluble carbohydrate in bulblets regenerated in vitro, with glucose and fructose present at significantly lower concentrations. Starch-degrading enzymes α- and β-amylase exhibited higher activities in both cultivars after cold treatment. Varying level of sucrose phosphate synthase, sucrose synthase and invertase activities were also observed after cold treatments.

신초 Cluster 형성에 의한 Lilium Asiatic Hybrid 'Hae Hwa'의 기내번식

Lilium asiatic hybrid 'Hae Hwa'의 인편 및 자구절편 배양으로 신초 cluster를 유도하여 자구를 대량증식하고, 형성된 신초 cluster에서 정상적인 자구를 대량생산하고자 일련의 실험을 실시하였다. MS배지에 BA 1.0mg/L와 IAA 0.5 mg/L가 첨가된 배지에서 신초 cluster의 유도는 인편보다 자구절편을 배양하는 것이 효과적이었다. MS배지에 BA 5.0 mg/L와 IAA 0.5 mg/L 첨가배지에서 신초수 및 신초무게가 양호하여 형성된 신초 cluster 절편에서 신초 cluster의 증식에 적합하였다. 또한 자구의 비대는 MS 배지에 sucrose 60∼90 g/L가 첨가된 배지에서 양호하였다. MS배지에 sucrose 30∼90 g/L를 주입한 5L airlift 생물반응기에 신초 cluster 절편체를 배양한 결과, 신초만 무성하게 자랐고 자구비대는 불량하였다. 신초 cluster를 8주간 배양한 후에 2배 MS 염류와 sucrose 250g/L, 활성탄 5 g/L를 포함한 액체배지를 동일용기에 첨가하여 배양한 결과, 명배양에서 형성된 자구수는 증가하였으나 소자구의 생장은 명, 암 모두 비슷하였다. 이로써 형성된 소자구의 무게가 53∼68 mg이며, BA 5.0 mg/L와 IAA 0.5 mg/L가 첨가된 MS 배지에서 신초 cluster를 증식한 후, 액체배지 첨가방법에 의하여 효율적인 소자구의 생산이 이루어졌다. This experiment was conducted to micropropagate bulblets via shoot cluster formation and massproduce normal bulblets from the sections of proliferated shoot clusters in Lilium asiatic hybrid 'Hae Hwa'. The induction of shoot clusters from the culture of bulblet sections was more effective than that of bulb scales on MS medium with 1.0 mg/L BA and 0.5 mg/L IAA. Proliferation of shoot clusters from the formed shoot cluster sections was the most favorable on medium containing 5.0 mg/L BA and 0.5 mg/L IAA. The formation and the growth of bulblets from shoot cluster sections were achieved effectively on medium with 60∼90 g/L sucrose. The leaves derived from shoot clusters grew vigorously but the bulblets from shoot clusters grew very poor in 5L air-lift bioreactor culture. By the addition of 30 mL fresh liquid medium containing doulble strength MS salts, 250 g/L sucrose and 5 g/L activated charcoal after 8 weeks in the shoot cluster culture on MS medium with 5.0 mg/L BA and 0.5 mg/L IAA, the number of bulblets was increased in light condition, but the growth of bulblets was not affected by light. Bulblet production was possible with the bulblet product at 53 to 68 mg in fresh weight by liquid medium addition after the proliferation of shoot cluster.

Molecular evidence that a lily-infecting strain of Tulip breaking virus from Japan is a strain of lily mottle virus

The sequence of the 3′-terminal 2074 nucleotides (nts), excluding the 3′-poly (A) tail, of RNA of a potyvirus isolated from lily (Lilium Asiatic hybrid cv. Enchantment) in Japan, currently tentatively designated as Tulip breaking virus-li (TBV-li), was determined. The sequence started within a single open reading frame (ORF) that encoded the carboxyl terminus of the large nuclear inclusion protein (NIb) and the complete 275-amino-acid coat protein (CP), followed by a 3′-untranslated region (3′-UTR) of 204 nts. The CP of TBV-li shared 91% amino acid (aa) sequence identity with that of TBV lily strain Dutch isolate (TBV-lily). The nt sequences of their 3′-UTR were 94% identical. However both viruses shared only 60–65% sequence identities with TBV tulip strain Niigata isolate in the corresponding regions. The results suggest that TBV-li is closely related to TBV-lily, and that these two TBV lily strains should be classified into a species different from TBV tulip strains. We therefore support a proposal to rename TBV-lily Lily mottle virus (LMoV), and suggest that TBV-li is another strain of LMoV (LMoV-J).

Postproduction Evaluations of Potted Asiatic and Oriental Hybrid Lilies

Potted Lilium Asiatic hybrids `Aristocrat', `Horizon', and `Polka' were evaluated following 3, 6, or 9 days of transport at 2, 7, or 13C. `Aristocrat' and `Horizon' withstood transport with little or no effect on floral bud opening. `Polka' was the most sensitive cultivar to transport, where bud opening decreased 33% when transported at 13C for 9 days. Most floral buds opened on `Aristocrat' (90% to 98%), while fewer buds opened on `Horizon' (37% to 56%) and `Polka' (52% to 90%). Individual flower longevity and diameters were largely unaffected by transport. Plant longevity was reduced 4 to 7 days when transported for 9 days at ≥7C or for >3 days at 13C. Plant longevity averaged 16 days for `Aristocrat' and `Polka' and 12 days for `Horizon'. `Aristocrat' and the Oriental potted hybrid lily `Star Gazer' were maintained at postproduction conditions of 18, 21, or 24C at 7 or 14 μmol·m–2·s–1 after being commercially transported for 4 days at 5 ± 2C. Postproduction conditions had no effect on floral bud opening of `Aristocrat' (98% to 99%), while bud opening of `Star Gazer' was reduced 17% at 24C compared to 18C. Plants lasted 4 and 9 days longer at 18C than at 21 or 24C, respectively. Foliar discoloration was greatest at 24C. Irradiance level had no effect on the variables evaluated.

ROLE OF PERLITE IN FLUORIDE TOXICITY OF FLORAL CROPS

Five common sources of perlite used in the North American horticultural industry were tested for their release of fluoride during five successive teachings with pH 5.2 sodium phosphate buffer (1 cc perlite/ml buffer). Soluble fluoride in the first leachate ranged from 0.05 to 0.8 mg/l and in the fifth leachate from 0.01 to 0.18 mg/l. Lilium Asiatic hybrids `Pixie Grange' and `Sunray', Chlorophytum comosum (Thumb.) Jacques `Variegatum', and Gibasis pellucida (Martens & Gal.) D. Hunt `Bridal Veil' were. grown in sphagnum peat moss plus perlite substrates ranging up to 50% perlite and at substrate pH levels from 4.6 to 7.0. No fluoride toxicity occurred in these crops in any treatments. The precaution against the use of perlite in substrates used for growing fluoride sensitive crops needs to be reconsidered.

The effect of sucrose, silver thiosulphate and 8-hydroxyquinoline citrate on the quality of Lilium inflorescences cut at the bud stage and stored at low temperature

A continuous supply of sucrose together with 8-hydroxyquinoline citrate to cut Lilium Asiatic hybrid ‘Prima’ inflorescences resulted in buds opening satisfactorily and increased their longevity. The best results were obtained using 30 g l −1 sucrose. Cut Lilium inflorescences could be stored at 1°C for 4 weeks without a great loss in potential vase-life and decorative value when the inflorescences were pre-treated with silver thiosulphate + 100 g l −1 sucrose for 24 h before cold storage, kept in a cold room in a solution containing 50 mg l −1 silver nitrate, and after cold storage kept in a solution containing 30 g l −1 sucrose and 200 mg l −1 8-hydroxyquinoline citrate. Such treatment greatly improved bud opening, increased the diameters of individual flowers and prolonged their life.

Scanning electron microscopy of meiotic chromosomes of plants in situ

Scanning electron microscope studies of anthers of the Asiatic Lilium hybrid Enchantment and Solanum tuberosum L. cv. Netted Gem fixed in organic acid – alcohol type fixatives clearly revealed nucleoli, bivalents, and the meiotic spindle. Centromere regions could not be identified in pachytene bivalents, but areas of possible spindle attachment were evident for metaphase I – anaphase I bivalents.