Analysis of plants obtained by embryo rescue from an interspecific Actinidia cross

The cross Actinidia chinensis Planch. var. chinensis (2x) × A. melanandra Franch. var. melanandra(4x) was made in 1989. Embryos were rescued early in seed development and raised in sterile culture. Plant material from these embryos was treated with colchicine to double the chromosome number and then put through a second shoot regeneration cycle. About 500 plants have been produced representing 25 clones from individual embryos. A sample of these plants has been analysed for chromosome number and for eight morphological characters. Plants with chromosome numbers representing 2x, 3x, 4x and 6x have been found and these numbers are correlated with some morphological characters. The origin of the different ploidy groups is discussed.     

In vitro induction of the amphiploid in interspecific hybrid of blueberry (Vaccinium corymbosum × Vaccinium ashei) with colchicine treatment

Amphiploids were produced from pentaploid hybrids between Vaccinium corymbosum (4x) and Vaccinium ashei (6x) by colchicine treatment. Seeds of each parental species and those obtained by the interspecific crossing were treated with colchicine at 0, 500, 1000 and 2000 mg/l for 7 days, and the ploidy level of the seedlings was determined with flow cytometry. Either amphiploid (10x) or ploidy chimera (5x + 10x) was obtained from interspecific hybrid seeds treated with all colchicine concentrations, while no chromosome-doubled plants were obtained from both parents. Cross direction in interspecific hybridization affected the results of colchicine treatment and amphiploids were obtained only when V. corymbosum was used as the seed parent. In this cross, 17–25% of the seedlings turned to be amphidiploids or ploidy chimeras by treating with 500 mg/l colchicines for 7 days. These results indicate that susceptibility to colchicine may increase in the interspecific hybrid compared with the parental species but only when V. corymbosum was used as female parent.     

Hormone effects on in vitro proliferation and rooting of Grevillea explants

Explants of Grevillea rosmarinifolia A. Cunn. and Grevillea×semperflorens F.E. Brigs ex Mullig., obtained from vegetative stem segments, were grown on a modified MS medium, which was supplemented with different contents of cytokinins (BA or K) to evaluate optimal conditions for bud proliferation. Auxins (NAA or IBA) were tested to evaluate rooting.

The multiplication rate was often higher in G. rosmarinifolia than in G.×semperflorens. Explant activity varied significantly depending on the cytokinin type and its concentration. The addition of cytokinins to the culture medium often promoted an increase in the number of shoots produced by each propagule, but at the highest BA concentration, shoot differentiation was inhibited in Grevillea×semperflorens. Only the higher concentrations studied (5–10 mg/l) produced better results with K, whereas BA was more effective at 1–2 mg/l. Rooting efficiency of the young shoots was lower in G. rosmarinifolia than in G.×semperflorens. IBA promoted root development in both species and was more effective at the highest concentration (1 mg/l). NAA gave better results at low concentrations (0.2 mg/l) in G. rosmarinifolia, while it had slight or negative effect on rooting of G.×semperflorens.     

Microsporogenesis and crossing behavior of a tetraploid, interspecific inter-EBN hybrid potato

The commercial potato Solanum tuberosum L. autotetraploid, has many wild relatives with the ploidy range of 2–6×, which are suitable sources of traits interesting in potato breeding. However, not all of these species can be readily crossed with cultivated potato due to pre- and post-zygotic barriers. Post-zygotic barrier on Solanum spp. is attributed to their difference in endosperm balance number (EBN). According to the EBN successful crosses need to have a 2:1 ratio of maternal to paternal EBN in hybrid endosperm. Thus, only parents with the same EBN can be crossed. The cultivated potato is 4EBN, while wild allotetraploid relatives such as S. stoloniferum are 2EBN. In this study, meiosis and crossability of an exceptional tetraploid 3EBN hybrid obtained from our previous research by crossing S. stoloniferum × S. tuberosum with 2 and 4EBN testers were studied. Although the exact chromosome pairing analysis was not possible, the formation of several polyvalents in each meiocyte suggests that recombination could occur between different genomes. Meiosis had many irregularities such as unequal segregation as well as having laggard chromosome and micronuclei. Crosses involving the interspecific hybrid, as the male parent with 2 and 4EBN testers were not successful, some of which were apparently due to the pollen–pistil incompatibility. Interestingly controlled self-pollination resulted in viable seeds. Crosses of the interspecific hybrid as pistillate parent in the absence of pre-zygotic barrier in most case were successful. Results of the study confirmed the segregation of EBN value in the gametes of hybrid. It was expected that backcross progeny of this hybrid would be a good source for selecting aneutetraploids.     

Morphological and cytological studies of diploid and colchicine-induced tetraploid lines of crape myrtle (Lagerstroemia indica L.)

The tips of cotyledon-stage seedlings of three crape myrtle cultivars (“Zi Wei”, “Hong Wei” and “Yin Wei”) were treated with colchicine. Various concentrations of colchicine and different treatment durations were tested. Seedlings of “Zi Wei” treated with 0.5% colchicine for 72 h and seedlings of “Yin Wei” treated with colchicine (0.2% for 96 h, 0.5% for 48 h and 0.8% for 72 h) demonstrated high rates of mutation; “Hong Wei” showed a slightly lower rate. The highest rate of morphological variation was 54.17% and this was achieved when tips were treated with 0.5% colchicine for 72 h. Putative tetraploid plants were identified with morphological and cytological variations, such as larger and thicker leaves, darker green coloration, larger stomata, lower density of stomata across the lower leaf epidermis and increased numbers of chloroplasts per stomata guard cell. Flow cytometric analysis demonstrated that the nuclear DNA content of 15 of these putative tetraploids (of which 7 were “Zi Wei”, 5 were “Yin Wei”, and 3 were “Hong Wei”) was indeed doubled relative to that of control diploid plants. The chromosome number of the tetraploid plants was 2n = 4x = 96, while that of the control diploid plants was 2n = 2x = 48. Ornamental characteristics were also enhanced in the tetraploid plants, with increased diameter of the individual flowers and greater basal length of the petals and claws. Pollen diameter and the size of capsules and seeds were also significantly greater than those of diploid plants.     

抗菌肽基因Gnk2-1经花粉管通道法导入西瓜的初步研究

 通过花粉管通道介导银杏抗菌肽基因Gnk2-1 于西瓜自交系‘04-1-2’中，以浓度为100、200、 300 和400 ng · μL-1 的DNA 为供体，分别于授粉后24、27、30 和33 h 导入，对子代进行PCR 检测。结 果表明：T0 代坐果率和单瓜结籽数随授粉后处理时间的增加而升高，但在各个处理时间下均显著低于对 照；外源DNA 的导入对授粉后27 h 处理的T1 代种子发芽率和授粉后24 h 处理的T1 代种子出苗率均造成 显著降低；不同DNA 浓度转化的T1 代种子的发芽率和出苗率与对照相比差异不显著；对1 280 株T1 幼 苗进行PCR 检测，得到32 株阳性转化植株，总体转化率为2.5%，最佳转化时间为授粉后24 ~ 27 h，最佳 DNA 转化浓度为100 ~ 200 ng · μL-1；PCR 阳性植株抗病鉴定表明转基因植株对西瓜枯萎病的抗性有所增强。     

In vitro propagation of Symphytum species

Methods are described for the in vitro propagation of Symphytum × uplandicum Nyman from bud, root and stem explants. Highest shoot numbers were produced from root explants > 4 mm in diameter, cultured vertically with their distal cut surface on the medium. The most suitable medium for shoot production was Murashige and Skoog's (MS) with 0.3 mg l⁻¹ 6-benzylaminopurine (BAP). These shoots developed roots on MS medium without hormones and were successfully transplanted into pots. Subculturing shoots onto MS medium containing BAP, kinetin (K), 6-λ,λ-(dimethylallylamino)-purine or gibberellic acid (GA₃) at 0.5, 1.0, 5.0, 10.0 or 30.0 mg l⁻¹ failed to stimulate outgrowth of axillary buds in culture. In vitro propagation from root explants was also achieved with S. asperum Lepech., S. officinale L. and S. × uplandicum cultivars ‘Bocking 1’, ‘Bocking 2’, ‘Bocking 4’ and ‘Bocking 17’, but not with S. bohemicum, S. grandiflorum DC, S. tuberosum L. or S. × uplandicum ‘Bocking 7’ and ‘Variegatum’.     

The growth and flowering of Hymenocallis × festalis

Bulbs of Hymenocallis (including Ismene) have showy, fragrant flowers. Little is known of the horticultural potential of these plants, and observations and trials on a stock of Hymenocallis × festalis are described.

In a stock of glasshouse-raised bulbs, bulb grade exerted a marked effect of the number of florets produced, which increased from 2 in 9–10-cm-circumference bulbs, to 7 in 18–19-cm-grade bulbs. Field-raised bulbs of the same grades produced fewer florets. Bulbs were usually planted for flower production in the glasshouse in April; earlier planting (February) leading to greater floret size but a longer period in the glasshouse before anthesis. Planting could be delayed at least until June, or later flowering could be achieved by storing the dry bulbs over winter at a low temperature (5°C), but the latter treatment reduced the percentage of bulbs which flowered. The long scapes could be dwarfed by ethephon (as Ethrel). Hot-water treatment, as a pest and disease control measure, did not result in damage to the flowers provided it was delayed until after the staminal cup initial had been formed.

In the field, growth of the main bulb was more vigorous than in the glasshouse, but pot culture in the glasshouse led to copious offset production. Data are presented for bulb increases for various grades of bulbs planted in outdoor beds at rates of 350–1050 g per metre row.     

Influence of sprayable mulch colour on yield of eggplant (Solanum melongena L. cv. Millionaire)

Planting beds were treated with sprayable, synthetic latex film (BASF, Charlotte, NC, USA) and oversprayed with black, white, red, blue, yellow or silver oil-based paint to study the effect of coloured spray mulches on yield of eggplant (Solanum melongena L. cv. Millionaire). Nonpainted mulch base and bare ground plots served as control treatments. Individual experimental units consisted of three rows, 102 cm wide × 7.3 m long, transplanted with 8 plants per row. Treatments were replicated in four blocks each at two separate locations near Fresno, CA. Plants grown on silver-painted beds produced significantly greater numbers (42–221%) of fruit and total fruit weight (42–237%) than all other treatments at both locations after eight harvests. Silver-painted mulch stimulated significantly greater flowering and fruit set numbers prior to the first harvest at one site. Apart from silver, only blue- and white-coloured mulches provided increased numbers and/or fresh weight yields at one site only. Mulched plots painted with other colours did not produce yields different from either the bare ground or nonpainted mulch base controls at either site.     

毛酸浆浸渍法多倍体诱导技术

以毛酸浆露白种子为试验材料,采用浸渍法进行多倍体诱导,研究了秋水仙素浓度和浸渍时间对后代表型变异率和死亡率的影响,以期筛选适合毛酸浆的多倍体诱导技术体系。结果表明:随着秋水仙素浓度升高和处理时间的延长,毛酸浆下胚轴膨大率升高,但过长的处理时间(36 h)会造成下胚轴膨大率下降。变异株气孔变大、密度变小,染色体数目鉴定结果为正常株2n=2x=24条,变异株2n=4x=48条。不同品种对秋水仙素浓度敏感程度不同,"铁把青"对秋水仙素较敏感,最适宜处理组合为100 mg·L^-1秋水仙素处理12 h,30 d表型变异率可达8.67%;"大黄菇娘果"和"粒粒甜小菇娘"最适宜处理组合为200 mg·L^-1秋水仙素处理12 h,30 d表型变异率分别为6.33%和6.67%。流式细胞仪鉴定发现很多表型变异株是混倍体,流式细胞仪可以在早期对变异株的倍性进行准确鉴定,还可以减少混倍体对育种的影响。     

A chilling-insensitive stage in germination of a low-temperature-adapted radish, rat’s tail radish (Raphanus sativus L.) cv. “Pakki-hood”

The rat’s tail radish (Raphanus sativus cv. “Pakki-hood”), cultivated in the northern part of Thailand, does not require low temperature for flower-bud initiation and shows high germinability even at low temperature. To analyze its characteristic temperature response, seedlings were subjected to a 3 day-chilling treatment at 0°C after a series of incubations for germination at 26°C which ranged from 3 h to 4 days. The chilling treatment after an initial stage (3 h to less than 2 days) of incubation at 26°C did not significantly affect the hypocotyl regrowth, but the same treatment after a more than 2-day incubation at 26°C reduced subsequent hypocotyl growth of plants. 1-aminocyclopropane-1-carboxylic acid (ACC)-dependent ethylene-forming activity was also reduced by the chilling treatment after 2–4-day incubation at 26°C. Electrolyte leakage from tissues of seedlings in the chilling treatment was increased in proportion to the length of the incubation at 26°C, indicating severe damage of chilling to plasma membranes after a longer incubation. We found a transition of chilling-sensitivity at 2 days in the incubation period for germinating seeds of Pakki-hood. Seedlings of Pakki-hood chilled earlier than the transition are relatively unaffected in their subsequent growth, but the seedlings become sensitive to chilling after the transition.     

Optimal pollination environment of tetraploid ginger (Zingiber officinale Roscoe) evaluated by in vitro pollen germination and pollen tube growth in styles

Pollen germination percentages in vitro of a tetraploid ginger (Zingiber officinale Roscoe), ‘4×Sanshu’, tended to be highest at around 20°C. Pollen tube growth in the styles was greatly enhanced at 17°C, i.e., pollen tubes penetrated into the entire stylar length in 66.7% of the styles used. Pollen stored for at least 3 h under 40–80% relative humidity (RH) almost completely lost its germinability, whereas pollen incubated for up to 3 h under 100% RH retained a relatively higher germinability. Moreover, the pollen germinability that was lowered under 40–80% RH did not recover when the pollen was re-humidified for 2 h under 100% RH before inoculation onto a medium. When stigmas were pollinated with fresh pollen at 17°C under 100% RH, pollen tubes penetrated into the entire stylar length in 26.7% of the styles tested. Under 40–80% RH, however, pollen tubes stopped growing at the middle or the base of styles. From these results, I concluded that the optimal pollination environment of the tetraploid ginger was at 17–20°C under 100% RH.     

Effect of Phaeomoniella chlamydospora and Phaeoacremonium sp. on in vitro grapevine plants

Artificial inoculation with fungal species involved in the Petri disease was performed on in vitro grapevine shoots. After 2 months, plantlets were observed and the presence of any leaf chlorosis or necrosis, considered as symptoms of the disease, were recorded. The fungal inoculation caused an increase in the number of symptomatic in vitro plants and in the incidence of external symptoms. Recovery of the fungal species was obtained after incubation of excised tissue fragments from the inoculated in vitro plants while some fertile fragments were collected also from plants without external symptoms. Higher levels of symptoms were observed in the rootstock varieties inoculated with Phaeomoniella chlamydospora while the lowest incidence was observed in the Aglianico cultivar. The possibility to correlate the visual assessment of in vitro plants with the data obtained by the analysis of digital images was confirmed.     

Genetic diversity and relationships of lotus (Nelumbo) cultivars based on allozyme and ISSR markers

Genetic diversity and genetic relationships of lotus (Nelumbo Adanson) cultivars were evaluated using allozyme and ISSR markers. The samples used covered 11 accessions of possible hybrids between Nelumbo nucifera and Nelumbo lutea and 92 accessions of N. nucifera including 69 flower lotus, 13 rhizome lotus, 5 seed lotus and 5 wild lotus. For allozyme studies, a total of 31 alleles at 23 loci of 18 enzyme systems were detected of which 5 (21.7%) loci Aat, Idh, Mdh-2, Pgd, Sod were polymorphic. The loci of Aat and Idh included two alleles, Mdh-2, Pgd and Sod included three alleles. Eighteen genotypes were detected with the 13 alleles of the 5 polymorphic loci. The parameters of average allele number, observed heterozygosity, expected heterozygosity and Shannon information index of 92 N. nucifera samples were 1.35 ± 0.71, 0.06 ± 0.21, 0.05 ± 0.14, 0.10 ± 023, respectively. Thirteen ISSR primers generated 93 loci, of which 37.63% were polymorphic across all samples. The percentage of polymorphic loci, average allele number, expected heterozygosity and Shannon information index of 92 N. nucifera samples were 26.67%, 1.30 ± 0.46, 0.10 ± 0.18 and 0.15 ± 0.25, respectively for the ISSR data. The ‘Bottleneck effect’ and rapid propagation of clones after the ice ages may explain the low genetic diversity of lotus. The dendrograms based on ISSR and allozymes were not congruent. Based on the ISSR data, the 103 samples were divided into the N. nucifera group (Group I), and the group containing inter-specific hybrids between N. nucifera and N. lutea (Group II). The flower lotus, rhizome lotus, and seed lotus each has multiple sources of origin. Plant size, a criterion commonly used in the classification of cultivars of lotus, is not correlated with genetic variation. Flower color is correlated with the cultivar classification to some degree, but its variation is complex in the hybrids.     

诱导金鱼草多倍体的初步研究

本文叙述了用秋水仙素水溶液处理金鱼草种子获得多倍体的方法，以及对获得的四倍体金鱼草（2n=32）与二倍体对照苗（2n=16)进行了形态、细胞学及气孔的观察比较。试验结果表明，浓度为0.3--0.5%的溶液处理24小时诱变效果较好，诱变率可达10--11.4%。镜检发现诱变植株的染色体多为混倍体，必须将混倍体形态明显似多倍体的部位进行扦插才可获得四倍体。四倍体金鱼草的形态从子叶期到大苗的各个发育阶段均与二倍体有明显差异，表现为叶子增厚、变形，气孔增大，花瓣增厚、增大，花期晚，自花授粉不孕等。四倍体植株的花粉因减数分裂异常，全部败育，是四倍体自花授粉不孕的主要原因。     

Exogenous polyamines alleviate the lipid peroxidation induced by cadmium chloride stress in Malus hupehensis Rehd.

In order to investigate the effects of spermidine (Spd) and spermine (Spm) on the stress of cadmium chloride (CdCl₂), it was checked that the content of polyamines and malondialdehyde (MDA), the rate of superoxide anion (O₂⁻) generation and the activities of superoxidase dismutase (SOD) and guaiacol peroxidase (GPX) in 1-year-old Malus hupehensis Rehd. var. pinyiensis Jiang (tea crabapple) seedlings grown in nutrient solution. The result showed that the content of free putrescine (Put) was increased, the contents of free Spd and Spm and the (Spd + Spm)/Put ratio in leaves were decreased significantly under CdCl₂ treatment. Foliar spraying with Spd or Spm under CdCl₂ treatment, the accumulation of free Put reduced, the contents of free Spd and Spm and the ratio of (Spd + Spm)/Put in leaves increased significantly. The activities of SOD and GPX in leaves were enhanced and O₂⁻ generation rate and MDA content were decreased significantly by foliar spraying with Spd or Spm under CdCl₂ treatment. Results suggest that both Spd and Spm can alleviate the lipid peroxidation caused by CdCl₂ in M. hupehensis var. pinyiensis.     

Colchicine- and trifluralin-mediated polyploidization of Rosa multiflora Thunb. var. inermis and Rosa roxburghii f. normalis

Rosa multiflora Thunb. var. inermis and Rosa roxburghii f. normalis are two important diploid species for rose breeding. Interspecific crosses between diploid species and tetraploid rose cultivars result in triploids with generally decreased fertility. A most promising gene transfer strategy is the production of fertile tetraploid plants before mating with tetraploids. In our study, germinating seeds of R. multiflora and R. roxburghii were treated with colchicine, and cotyledon-stage seedlings of R. multiflora were treated with colchicine or trifluralin in order to obtain tetraploid plants. Also, various concentrations of antimitotic agents and different treatment durations were tested. The results showed the antimitotic agents affected the plant morphological characteristics (i.e. plant height, stem diameter) of R. multiflora and R. roxburghii. Twenty-three tetraploid plants were obtained from the germinating seed treatments (21 from R. multiflora and two from R. roxburghii), but no tetraploid plants were obtained from the seedling treatments. The highest rate of chromosome doubling (25.0%) was achieved when germinating seeds of R. multiflora were treated with 0.2% colchicine for 12 h. At the same time, marked morphological variation was observed and analyzed among tetraploid plants and their corresponding diploids of R. multiflora.     

Optimizing the lighting regime for Alstroemeria with respect to photoperiod and fluence rates

The Alstroemeria cultivars Diamond, King Cardinal and Libelle were grown for 18 months under five lighting regimes with, and without, soil cooling. The aim was to optimize the daily investment of light energy from artificial sources with respect to photoperiod and photosynthetic fluence rates and to elucidate possible links between reactions to photoperiod and root-zone temperature. The more photons (photosynthetically active radiation, PAR) that were supplied to the plants per day (8, 11 and 13 mol m⁻²), the higher was the total production of flowering stems. The total yield from regimes with 13 mol m⁻² day⁻¹ was higher when the light was spread over 20 and 16 h compared to 12 h. In treatments with soil cooling, the plants flowered continuously under all combinations of photoperiods and photosynthetic fluence rates, and the summer and autumn recession in flower production that occurred for non-cooled ‘King Cardinal' and ‘Diamond' was the same under all lighting regimes. It is concluded that it might be more cost-effective to spread the daily investment of light over 20 rather than 16 or 12 h when the total energy budget and CO₂ costs are taken into consideration.     

Induction of tetraploidy in meristematically active seeds of Japanese barberry (Berberis thunbergii var. atropurpurea) through exposure to colchicine and oryzalin

Japanese barberry (Berberis thunbergii DC) is an invasive shrub, widely naturalized across the United States, whose numerous cultivars remain an important horticultural commodity. Maintaining this crop for the future necessitates the development of sterile clones. Exposure to the mitotic inhibitors colchicine and oryzalin is a traditional method for inducing tetraploidy in breeding lines as a precursor to creating sterile genotypes. Treatments utilized pre-germinated B. t. var. atropurpurea seeds with emerged radicles. Seeds were immersed in aqueous solutions of colchicine (.02%, .05%, .1% and .2%) and oryzalin (.002%, .005%, .01% and .02%) dissolved in 1% DMSO for 6, 12 and 24 h durations. Seedling ploidy level was determined via flow cytometry following 6 and 52 weeks of growth in the greenhouse. Both anti-mitotic chemicals proved effective at inducing tetraploidy and produced comparable efficiency rates. The survival rate of treated seeds decreased in response to both increased mitotic inhibitor concentration and longer exposure duration. While exposure to oryzalin produced greater seed mortality than colchicine, most seedlings that survived had altered ploidy levels. The most efficient oryzalin concentration was 0.002% with a rating of 28%, while the most efficient colchicine concentrations were in the range from 0.05% to 0.2%. Duration of exposure to mitotic inhibitor was not a significant factor over the range from 6 to 24 h. Reversion of tetraploid plants to the diploid state occurred at a low frequency following a dormancy period. Some tetraploid seedlings derived from exposure to both chemicals displayed foliar abnormalities including irregular leaf margins and mottled lamina. The primary advantage of colchicine was low seedling toxicity, while oryzalin was notable for its ability to induce tetraploidy at low concentrations.     

Measurement of soil compaction tolerance of Lagestromia speciosa (L.) Pers. using chlorophyll fluorescence

Trees planted along roadsides and on public recreation areas are subjected to environmental stresses such as poor soil, air pollution and heat. However, very little information is available on the trees’ tolerance to the various stress factors faced in an urban environment in Malaysia, such as soil compaction. The effects of soil compaction on a range of chlorophyll fluorescence parameters (F₀,F_m,F_v/F_m) in foliar tissue of Lagestromia speciosa, a widely planted Malaysian street tree, were examined. Results showed that soil compaction was between 170 and 315 MPa in the study areas. Soil compaction above 180 MPa affected tree form and reduced chlorophyll fluorescence. It is concluded that chlorophyll fluorescence offers a rapid screening technique for assessing soil compaction tolerance of L. speciosa.