A thermal time model for predicting time to aerial shoot elongation in Variegated Solomon's Seal

Aerial shoot development in Variegated Solomon's Seal (Polygonatum odoratum‘Variegatum’) was studied under warm (mean 18°C) conditions after dormant rhizomes had been stored at a range of temperatures. After chilling at 0.8–5.5°C for 21–77 days, all rhizomes produced elongated aerial shoots, with mean lengths from 33 cm to 44 cm. Exposure of rhizomes to 15°C or 20°C for 21–77 days resulted in 17% to 50% of buds emerging as shoots, but these either aborted or failed to extend beyond a rosette. The earliest aerial shoot elongation was observed after 7–13 days at 18°C in rhizomes that had been chilled at 0.8–2°C treatments for 59–72 days. The base, optimum and maximum temperatures during pre‐planting storage for subsequent aerial shoot elongation were derived respectively as ?1.5°C, 1.9°C and 15.8°C. A thermal time of storage was calculated from these cardinal temperatures and the rate of progress to normal aerial shoot clongation was shown to increase linearly with increasing thermal time to c. 150°Cd. The thermal time procedure for predicting time to aerial shoot elongation constructed from growth room/chamber data was validated using rhizomes that had been exposed to varying temperatures in shadehouse conditions during the winter in Taiwan.     

In vitro plant regeneration of Mammillaria elongata normal and cristate forms

In the normal form of Mammillaria elongata, shoots were regenerated in vitro, through callus, from tubercle explants excised from the upper part of the branch and cultured on Murashige and Skoog medium (MS) with 1.07 μM α-napthaleneacetic acid (NAA) and 22.20 μM 6-benzylaminopurine (BA). A high percentage of tubercles explants of the M. elongata cristate form, excised from the tip of the branch and cultured on MS with 0.54 μM NAA and 0.44 μM BA or 1.07 μM NAA, responded by initially forming an inflated cristate shoot, which gave cristate and normal shoots, without callus intervention, when transferred on basal MS. Callus formed on cristate tubercles explants gave both cristate and normal shoots when transferred onto basal MS. Normal and cristate shoots were rooted in vitro on MS with 9.84 μM or 0.98 μM indole-3-butyric acid, respectively, and established ex vitro. In both normal and cristate form, the differential response appeared to be associated with the site of the explant excision. The formation of shoots was influenced by the season of culture; i.e., explants excised in October had a higher shoot formation rate than those excised February. This revised version was published online in June 2006 with corrections to the Cover Date.     

Critical oxygen pressure for growth and respiration of excised and intact roots

A method based on the measurement of ATP/ADP ratios is described. It permits the determination of the critical respiratory oxygen pressure of any organ, or part of any organ, of an intact plant. The data obtained by this method with intact maize (Zea mays L. INRA 508) root tips are compared with polarographic determinations on similar excised tissues.

When internal O₂ transport from the aerial part was prevented, the critical oxygen pressure found for the respiration of intact tips was similar to that found with excised tips. It was close to 10 kilopascals in a humid atmosphere and about 30 kilopascals in a liquid medium. Flooding of the gas spaces by vacuum infiltration did not modify these results. When internal O₂ transport from the aerial parts of the plant occurred, significantly lower values were obtained in liquid medium for the critical oxygen pressure, which shifted from more than 21 to 6 kilopascals. The higher values observed with excised root tips, compared to those obtained with intact tissues, can be explained by the lack of internal O₂ transport, rather than by the flooding of gas spaces.

Data are presented which show that root growth started to be limited at a significantly higher pressure than the respiration. These results are attributed to nonrespiratory oxidative processes with a low affinity for O₂ involved in root elongation.

     

Composition of Excised and Attached Sterile Pea Roots--Loss of Cell Nucleic Acids

The composition of excised cultured pea-root tips has been comparedwith aseptically cultured attached root tips. Expressed in cellularterms, the cultured excised roots showed a reduction in nitrogenousand nucleic acid constituents. The cells of attached roots hadapproximately 2-, 4-, 5- and 4-fold amounts of protein, solublenitrogen, ribonucleic acid and deoxyribo-nucleic acid respectively,as compared with excised roots cultured in the same medium.Cultured root meristems did not contain the quiescent zone normallyfound in attached roots. It is suggested that the observed reductionin RNA and DNA per cell does not impair the ability of cellsto divide and expand normally and, further, that there is surplusnucleic acid in attached root cells not obligatory for fullexpression of the genome.     

In vitro micropropagation of Ruscus aculeatus

We have developed three protocols for the rapid micropropagation of Ruscus aculeatus. The primary explants utilised were immature embryos, aerial buds excised from rhizomes and shoot buds regenerated from organogenic calli. In order to increase the plant regeneration from the primary explants, we used organogenic calli from cladode, stem and rhizome segments. We tested more than 20 culture media for callus induction and shoot regeneration and the best results were obtained when rhizome segments were cultured on Murashige and Skoog medium supplemented with 0.5 mg dm⁻³ 2,4-dichlorophenoxyacetic acid and 1 mg dm⁻³ kinetin.     

The distribution of hop latent viroid within plants of Humulus lupulus and attempts to obtain viroid-free plants

The detectability of hop latent viroid (HLVd) was investigated in field-grown hop (Humulus lupulus L.; an herbaceous perennial in which all the aerial parts die at the onset of winter) plants, using dot-blot hybridisation. The viroid was readily detected in all aerial tissues in the second half of the growing season but it could not be detected very early in the season. Between early- and mid-season, HLVd was first detected at the base of the new stems and then apparently spread up them as they grew but only became detectable near the tips of the shoots at mid-season, approximately at the time most elongation growth ended and flowering began. Petioles were the most convenient tissues to test, being easy to collect and, relative to leaf lamina tissue, low in inhibitors. Both dot-blot and in situ hybridisation failed to detect HLVd in shoot tips taken from plants grown at two ‘low’ temperatures (10°C and 15°C). Failure to produce any viroid-free plants by in vitro culture from such tips suggested that they did contain viroid but at levels too low to detect by either method. Lower temperatures and smaller explants are now being investigated as means of producing viroid-free plants.     

Cryopreservation of persimmon (Diospyros kaki Thunb.) by vitrification of dormant shoot tips

Shoot tips excised from dormant axillary buds of persimmon (Diospyros kaki Thunb.) were cryopreserved by vitrification. These excised shoot tips were dehydrated in a highly concentrated vitrification solution for 20 min at 25°C and then plunged directly into liquid nitrogen. After rapid warming in water at 40°C, the shoot tips were rinsed in a 1.2 M sucrose solution for 20 min and then plated on a solidified culture medium. Successfully vitrified shoot tips resumed growth within 10 days of plating and developed shoots within 3 weeks without intermediary callus formation. This simple protocol was successfully applied to the 16 cultivars found in the temperate zone. The average rate of shoot formation was 89%. Even the subtropical species of Diospyros demonstrated a very high recovery growth when the shoot tips had been previously osmoprotected with a mixture of 2 M glycerol plus 0.4 M sucrose for 20 min following sucrose preculture. Little or no contamination occurred in the cryopreserved shoot tips excised from sterilized winter axillary buds. Thus, this simple and reliable vitrification protocol using dormant shoot tips appears to be promising as a routine method for the long-term conservation of Diospyros germplasm of both temperate and subtropical origins.     

Rhizome development ofPhragmites australis in a reed community

Variations in the height, shoot density, biomasses of above- and below-ground parts and rhizome distributions ofPhragmites australis were investigated along a line-transect in a reed community at Yufutsu Mire, Hokkaido. Relationships of performance of the reed plants to soil conditions and species compositions were also examined. Three types of rhizome development were recognized in reed plants; (1) the central part of the reed community, characterized by well developed rhizomes and dense aerial shoots, (2) the intermediate part, characterized by development of rhizomes along both the peat and surface layers and very dense aerial shoots, and (3) the marginal part, characterized by development of rhizomes only along the peat layer and sparse aerial shoots. Observation showed that rhizomes in the surface layer actively produced aerial shoots, whereas rhizomes in the peat layer contributed to the spreading of their distribution range. With the growth of rhizomes, organic debris originating from dead rhizomes and roots accumulated in the mineral soil to promote organic soil formation. In dense parts of the reed stand, species composition was poor because of the shading and litter accumulation by reed plants. The effects of microtopography and water level on the establishment of reed seedlings were also considered.     

爬树龙的组织培养

爬树龙(Rhaphidophora decursiva (Roxb.)Schott),别名过山龙,系天南星科(Araceae)崖角藤属(Rhaphidophora)的一种藤本植物,以气根攀缘于它物上生长,叶表面绿色、发亮,它不仅是一种多年生常绿的观赏植物,而且其茎叶入药,可医治许多病症,因此有一定的药用价值。通常用扦插方法繁殖,我们取它的茎尖和茎节进行组织培养试验,现将初步试验结果简要报道如下。     

Effects of post-chilling temperature on growth and variegation in Solomon's Seal

Effects of temperature were studied on the current and following season's growth of shoots from chilled rhizomes of Variegated Solomon's Seal. The rate of progress to completed elongation of the aerial shoot in chilled plants increased linearly with increasing temperature up to 28°C (24 h mean). A post‐chilling thermal time of 658 ± 47°Cd (> ‐1.3°C) was required for aerial shoots to become fully extended. Temperatures of 28°C and 33°C accelerated aerial shoot senescence and decreased rhizome and root dry weights, as compared with 18°C and 23°C treatments. Leaf number and variegation were not affected by temperature treatments during current growth season and all plants produced 12–13 leaves with between 7% and 9% leaf area variegated. Leaf variegation, however, was significantly increased in plants that had been grown after chilling at 28°C during the preceding growing season. Proteins of approximately 26, 32 and 62 kDa were present in the green parts of leaves but not in the white parts.     

Direct somatic embryogenesis and encapsulation of somatic embryos for in vitro conservation of Bacopa monnieri (L.) Wettst

Direct somatic embryogenesis and shoot organogenesis were achieved from leaf explants excised from microshoots of Bacopa monnieri cultured on Murashige and Skoog medium containing N6-benzyladenine (BA) and 2,4-dichlorophenoxyacetic acid (2,4-D). The maximum frequency of explants differentiated somatic embryos and shoot buds on MS medium supplemented with 12.5 µM BA and 1 µM 2,4-D. The frequency of explants differentiating somatic embryos decreased with increasing concentration of 2,4-D. Light and scanning electron microscopy revealed direct differentiation of somatic embryos and shoot buds from explants, and various developmental stages of the somatic embryos were observed. Somatic embryos and apical shoot tips were encapsulated in sodium alginate gel to produce synthetic seeds. The storage of synthetic seeds produced by encapsulation was studied at 4 and 25?°C (room temperature) for a period of 140 days. Encapsulated somatic embryos were found to retain viability after 140 days of storage at both temperatures, whereas encapsulated apical shoot buds failed to germinate even after 40 days when stored at 4?°C. The viability of synthetic seeds was higher when stored at 25?°C. All amplified markers scored by random amplified polymorphic DNA (RAPD) and inter-simple sequence repeats (ISSR) were monomorphic for all the plants produced from synthetic seeds following different periods of storage, thus establishing the clonal fidelity of propagated plantlets.     

SOME EFFECTS OF GAMMA IRRADIATION ON SEEDS AND RHIZOMES OF MUSA

The tolerance of Musa balbisiana Colla seeds to gamma irradiation was found to be considerably greater than that of rhizomes of the parthenocarpic variety ‘Gros Michel': e.g., 11.8 krad reduced the germination of rhizomes 92% and of seeds approximately 15%. Intact seeds exposed to doses higher than 48 krad did not germinate in non-sterile soil, but, when scarified and cultured axenically after irradiation, seeds which received doses as high as 70 krad germinated. Embryos excised from seeds exposed to doses as high as 285 krad formed callus, indicating that not all metabolic processes were inhibited by these extremely high doses. There was considerable variation in radiation tolerance between seed lots which was not related to their age, moisture content, or pre-exposure viability. Germination of intact seeds appeared to be stimulated by doses of 3 or 9 krad. No lasting differences attributable to the level of irradiation were apparent in the development of seedlings derived from either intact or scarified seeds nor of plantlets derived from excised embryos. Conversely, there was a significant reduction, proportional to irradiation dose, in the growth of plants developing from rhizomes, emphasizing the greater radiation sensitivity of the vegetative propagule. The radiation tolerance of seed-borne microorganisms was considerably higher than that of the plant materials, indicating that gamma irradiation is not effective as a means of obtaining pathogen-free rhizomes or surface-sterilizing seeds of M. balbisiana.     

Gibberellin-like Substances Obtained from Chilled Plants of Dianthus barbatus L. by an Agar Diffusion Technique

Plants of Dianthus barbatus with a cold requirement for floweringwere subjected to chilling treatments at 5 °C. An agar diffusiontechnique was used to collect gibberellin-like substances fromshoot tips excised from these plants and from plants that hadbeen kept in the glasshouse at a minimum temperature of 14 °C.Shoot tips from chilled plants gave markedly higher yields ofgibberellin-like substances. The effect of chilling was no longerso apparent if plants were returned to the higher temperaturesof the glasshouse for 1 week before the shoot tips were excised. The proportion of plants that flowered as a result of the differentchilling treatments varied widely but this variation was notassociated with any obvious differences in the yields of gibberellin-likesubstances. Application of gibberellins to plants grown at aminimum temperature of 14 °C did not promote flowering.     

In vitro production of Indian citrus ringspot virus (ICRSV) free Kinnow plants employing thermotherapy coupled with shoot tip grafting

Indian citrus ringspot virus (ICRSV) is known to cause serious disease problem in Kinnow (Citrus nobilis Lour × C. deliciosa Tenora) plants. This work reports the elimination of ICRSV by using thermotherapy coupled with shoot tip grafting in vitro. Nodal segments from infected mother plants (indexed by indirect ELISA and RT-PCR) were treated both in water bath and moist hot air at different temperatures viz. 40, 45 and 50°C for 30, 60 and 120 min and cultured on MS medium containing 2-iP (1 mg/l) and malt extract (800 mg/l). Shoot tips were excised from the nodal sprouts and grafted on to rough lemon (C. jambhiri) rootstock under aseptic conditions. Water bath treatment was found to be more effective as compared to moist hot air treatment as maximum number of ICRSV free plants (36.84%) were obtained by grafting the tips (0.7 mm) taken from the nodal segments treated at 50°C in water bath for 2 h. In an alternate treatment regime, 1-year-old infected plants were kept at various temperatures viz.36, 38 and 40°C in a thermotherapy chamber. Maximum of 60% ICRSV free plants were obtained by grafting the tips (0.7 mm) from the plants placed at 40°C followed by the plants placed at 38°C (59.09%) and the least was observed in case of the plants placed at 36°C (40.74%). Only those plants/plantlets were considered virus free, which showed negative reaction both in Indirect ELISA and RT-PCR.     

Effects of insulin on rat brain noradrenaline

A single intracardial injection of streptozotocin produced a significant increase in rat hypothalamic noradrenaline while no changes were observed in the olfactory tubercles. The parenteral administration of a single dose of insulin decreased rat hypothalamic noradrenaline; the effect had a rapid onset and lasted for at least six hours. Similar noradrenaline reductions were observed in the olfactory tubercles but in this tissue the depletion started later and recovered earlier. In addition, in olfactory tubercles after insulin injection, tyrosine level and dopamine metabolism were increased. The results show that the increases in hypothalamic NA observed in streptozotocin diabetic rats are counteracted by insulin administration and possibly the consequence of changes in noradrenaline turnover.     

莲的生态解剖学研究

莲叶有三种形式:沉水叶无气孔;浮水叶气孔仅分布于腹面;挺水叶形大、气孔数多。器官中气室和气道发达。排水器位于叶腹面的中央,除常敞开着外,与气孔相似,水分可从此排出。莲具有星散的维管束、无形成层,只在根中有导管,胚根简化,不定根发达。花早期发育在水中,后由柄生长延伸出水面。果实中具有三个功能不同的孔。果皮上的表皮层及栅状细胞层发达。     

In vitro loss of nucleic acids from cells of aseptically cultured excised pea roots

Summary Analysis of pea root tips taken from attached seedling roots and excised roots cultured in vitro has revealed major differences in cell constituents. The cells of cultured roots have only 40% and 13% of the protein and amino acid content of attached root cells. The nucleic acid content of cultured root cells was shown to be only 20% and 27% of the RNA and DNA respectively found in attached roots. It is suggested that there is excess nucleic acid in whole plant tissues above that required for transfer of genetic information necessary for normal growth and differentiation of root cells.     

Agrobacterium rhizogenes-mediated transformation and plant regeneration of four Gentiana species

Shoots of micropropagated Gentiana acaulis, G. cruciata, G. lutea, and G. purpurea were inoculated with suspensions of Agrobacterium rhizogenes cells, strains ATCC 15834 or A4M70GUS. Adventitious roots appeared at the sites of inoculation in all 4 species. Root tips were excised and cultured on growth regulator-free media for 2-6 years. They exhibited very high branching and plagiotropism. Spontaneous bud initiation occurred in roots of G. cruciata. Roots of G. lutea, G. acaulis and G. purpurea were cultured on media with high kinetin concentration, which induced the formation of friable callus tissues. Only in G. purpurea were these calluses organogenic. Regenerated shoots of G. cruciata and G. purpurea gave rise to plants, that displayed the typical phenotypes of A. rhizogenes-transformed plants: short internodes and rolled leaves. In the roots of G. acaulis and G. cruciata, transformed with A. rhizogenes A4M70GUS, a positive reaction with X-gluc indicated the activity of β-glucuronidase. The DNA extracted from hairy roots and from the roots of transgenic plants hybridized with the appropriate genomic probes in Southern blotting. This is taken as evidence of the stable genetic transformation in the 4 Gentiana species. This revised version was published online in June 2006 with corrections to the Cover Date.     

Cultivation of Excised Anthers in vitro--Effect of Nucleic Acids

Anthers of Allium cepa and Rhoeo discolor, excised at leptotene-zygoteneand diplotene-diakinesis, have been cultured in modified White'smedium supplemented with different concentrations of ribonucleicacid and desoxyribonucleic acid. The best development was obtainedwith ribonucleric acid in which there was a 100 per cent. survivalof the microspore mother cells up to the formation of 1-celledmicrospores. In R. discolor, the time taken by the microsporemother cells to form tetrads in vitri was half of that requiredin nature. All attempts to induce division of the microsporenucleus in vitro failed.     

The elimination of ryegrass mosaic virus from Lolium multiflorum by meristem tip culture

Meristem tips were cultured from Lolium multiflorum plants infected with ryegrass mosaic virus (RMV). Meristem tips within the size range O'2-fi mm long were cultured on media with or without 2 ,4-D at 1 mg/1. The plants that regenerated in culture showed no symptoms over a 10 month period and no RMV particles were observed by electron microscopy. It was concluded that RMV had been eliminated. The method should prove useful in eliminating the virus from desirable genotypes used as parent material for seed production.