May 1995, Volume 37 Issue 5


          Research Articles
Isolation of Pollen Protoplasts in Nicotiana tabacum
Author: Wang Yong and Zhou Chang
Journal of Integrative Plant Biology 1995 37(5)
    A new method for isolation of quantities of mature pollen protoplasts in Nicotiana tabacum has been established. The first step was to germinate mature pollen in Brewbaker and Kwack medium containing 20% sucrose. When most of the pollen grains had just germinated short pollen tubes, they were transferred to an enzymatic solution for the second step. The enzymatic solution contained 1% pectinase, 1% cellulase, 0.5% potassium dextran sulfate, 1 mol/L mannitol, 0.4 mol/L sorbitol in Dx medium with or without 15% Ficoll. The enzymes firstly degraded the pollen tube wall and then the intine. As a result, intact pollen protoplasts were released with the isolation rate up to 50%-70%. Factors affecting pollen protoplast isolation during the germination and maceration of pollen grains were studied. The suceees depended on two key points:pollen germination duration and osmotieum concentration. The optimal germination duration was 30 rain at 30. When it was too long, long pollen tubes formed and subsequently, large number of subprotoplasts instead of whole protoplasts were yielded, as the case reported by previous investigators. The optimal concentration of mannitol and sorbitol in enzyme solution was as high as 1.4 mol/L in total. Lowering of the osmoticum concentration resulted in decrease of percentage of pollen protoplasts.
Abstract (Browse 1738)  |  Full Text PDF       
Transgenic Rice Plants Produced by PEG-Mediated Plasmid Uptake into Protoplasts
Author: Li Wen-bin, Wang Ge-jiao, Stanchina E, Castiglione S, Sun Yong-ru and Sala F
Journal of Integrative Plant Biology 1995 37(5)
    Embryogenic cell suspension cultures were obtained from calli developed from mature rice seeds of a Japonica type Itahan cultivar, Roncarolo. Protoplasts were isolated and transformed by PEG-treated method with plasmid pHP23 carrying the NPT gene which encodes resistance to antibiotic G-418. Protoplast-derived colonies were selected in presence of the inhibitor. Plants were regenerated and transplanted into soil in the green house. The presence of foreign gene in the regenerated plants was verified by PCR and Southern analysis.
Abstract (Browse 1773)  |  Full Text PDF       
Observation of One Pheophytin a Molecule Not Associated with Primary Photochemistry in the Isolated Photosystem Reaction Center D1/D2/Cyt b559 Complex
Author: Hou Jian-min, Peng De-chuan, Kuang Ting-yun,Tang Chong-qin and Tang Pei-song
Journal of Integrative Plant Biology 1995 37(5)
    Photodamage of pheophytin a (pheo a) in the isolated photosystem (PS ) reaction center D1/D2/Cyt b559 complex from spinach has been investigated by high performance liquid chromatographic method in detail. The results showed that: (1) There is one pheo a molecule which is not associated with the primary photochemistry in the PS reaction center complex. It may be considered that there are two different electron transfer branches in the PS reaction center just as in the purple bacterium photosynthetic reaction center. (2) The damaged pheo a may be attributed to the one bonding to the D2 protein comparing the D2 subunit in the PS reaction center with M subunit in the purple bacterium photosynthetic reaction center. (3) A possible arrangement model of redox cofactors in the PS reaction center was proposed based on our experiment.
Abstract (Browse 1655)  |  Full Text PDF       
Study on Organelle DNA Within the Generative Cell and Sperm Cells in Pharbitis
Author: Hu Zan-min and Hu Shi-yi
Journal of Integrative Plant Biology 1995 37(5)
    The organelle DNA in generative cell and its behavior during spermatogenesis in Pharbitis limbata and P. purpurea were observed by epifluorescence microscopy stained with 4',-6-diamidino-2-phenylindole (DAPI). In these two species, the generative cell is long and thin in which a great amount of cytoplasmic DNA is present. Most pairs of sperm cells are isomorphic, in which one end is obtuse and the other is elongate, but in a few pairs dimorphi sperms are present. The nucleus is located at one end of the cell. A lot of cytoplasmic DNA are distributed randomly throughout the cytoplasm. The size of organelle nucleoids and their fluorescence intensity are different in a sperm cell. The features of generative cell and sperm cell, and behavior of cytoplasmic DNA are similar in P. limbata and P. purpurea. The obvious differences between them are that the size and fluorescence intensity of organelle nucleoids in P. purpurea are respectively smaller and weaker than in P. limbata. The results showed that morning glory has potential of biparental or paternal cytoplasmic in heritance. Isomorphism and dimorphism of sperms, and the relationship between the ratio of nucleus and cytoplasm in sperm cell and the plastid biparental inheritance are discussed.
Abstract (Browse 1628)  |  Full Text PDF       
Formation of Interfamilial Cell Symplast During Cell Co-Culture in Vitro
Author: Guo Xiao-cai, Wu Bo-ji, Jiang Hui and Zheng Guo-chang
Journal of Integrative Plant Biology 1995 37(5)
    Two cell lines, white tobacco (Nicotiana tabacum) cell line and green carrot (Daucus carota) cell line, each with very distinct cellular structure markers by which the two cell lines could be identified at levels of callus cells, with light microscopy and electron microscopy, were established and used for interfamilial cell co-culture in which callus cells were well mixed and finely dispersed and treated with K+ hypotonic solution. Variegated interfamilial chimeral calli were observed after 10 to 15 days of co-culture. An isolation layer was formed and became thickened at the interface between the two attached unrelated callus cells and the heteroplastic cell wall complex was gradually established. Then the isolation layer became thinned and disappeared and plasmodesmata were formed secondarily within the thinned region and the interfamilial cell symplast was established. The wall in the region with isolation layer was about twice as thick as the fused cell wall of the symplastic. The developmental process of the interfamilial cell symplastic connection was discussed and it was suggested that (1) the nonspecific formation of isolation layer as initial adhesion between the two attached unrelated cells was the prerequisit for symplastic connection de novo formation, and (2) the specific cell recognition leading to disappearance or thickening to lignification or suberization of isolation layer and formation of plasmodesmata started within the isolation layer.
Abstract (Browse 1833)  |  Full Text PDF       
Fluorescence Microscopic Observations on the Distribution Patterns of Actin Filaments in the Cells of Developing Endosperm in Triticum aestivum
Author: Zhang Wei-cheng, Jin Shi-ping and Yan Wen-mei
Journal of Integrative Plant Biology 1995 37(5)
    The presence and distribution patterns of actin filaments (AFs) in the cells of developing wheat (Triticum aestivum L. ) endosperm exhibiting intercellular protoplasmic movement were studied with fluorescence microscopy and video microscopy. By using TRITC-PhaIloidin as fluorescence probe and cytochalasin B (CB) treatment it was uncovered that there were a lot of AFs scattered throughout the cytoplasm and the patterns of AFs varied greatly with the actin localization. Four configurations of AFs could be recognized: an actin meshwork surrouding the nucleus; bundles of AFs radiating from nuclear basket and extending to the periplasm; numerous finer AFs densely and randomly distributed in the cortical cytoplasm and fusiform bodies composed of AFs appearing in the endosperm cells lying at the cheek of the caryopsis. Judging from the dynamic characters of intercellular movement of the cytoplasmic constituents and the reaction of cytoplasmic strands related to CB treatment, the authors have discussed and proposed that the exhibition of the two kinds of intercellular movement (extrusion of cytoplasmic strands and mass flow of ground substance) might also be in close relation to the different configurations of AF organization in the cytoplasm.
Abstract (Browse 1710)  |  Full Text PDF       
Biodiversity Studies on Quercus ilex Woods in Veneto, Italy
Author: Xie Jin-yang, Chen Ling-zhi, Ghirelli L and Chiesura Lorenzoni F
Journal of Integrative Plant Biology 1995 37(5)
    The species diversity indices and the species abundance distribution of Ouercus ilex L. woods in Veneto, Italy were studied. The results are as follows: 1. The Menhinicht s index, Shannont s index, Brillouins index, Simpsont s index Piulout s evenness, V evenness of PIE, V evenness of PIE, Mclntosh evenness could be used to study the diversity of O. ilex woods in Veneto. They showed that from the area of Lago ki Garda, to Foci del Tagliamento, Bosco Nordio, and Colli Euganei, the value of species diversity indices decreased gadually. 2. The geometric series of species abundance distribution dould be used best to study the diversity of O. ilex woods, and the log-normal distribution of species abundance distribution could also be a good index used to study did. The results of the study of species abundance distribution were the same as those of species diversity indices. 3. Diversity did not always increase in succession. In the earlier stage of succession, the diversity usually was relatively large owing to the invasion of many species and the lack of dominance. In the subsequent stage, the diversity was relatively minimized as the dominance increases. In the late stage, the diversity would increase as the structure of community became complicated and the number of species increased.
Abstract (Browse 2067)  |  Full Text PDF       
Plants Regenerated from Mesophyll Protoplasts of Cottonwood New Clone
Author: Wang Ying, Huang Min-ren, Chen Dao-ming, Xu Nong, Wei Zhi-ming and Xu Zhi-hong
Journal of Integrative Plant Biology 1995 37(5)
    Poplar NL-80106 (Populus deltoidesP, simonii) mesophyll protoplasts were isolated from leaves of 30 days-old sterile shoot, with 4 107/g fr. wt of protoplast yield after purification. The protoplasts were cultured in KM8p and MS liquid media containing 2 mg/L 2, 4-D, 0. 5 mg/L NAA and 0.5 mg/L KT. Higher plating density and lower osmatic pressure (0.45 mol/L) were proved to be favourable to division of protoplast-derived cells. The first division initiated 5 days after culture, and the division frequency reached 4.5 % on the 10th day. A number'of cell colonies and microcalli was formed in 12 weeks. Using organic nitrates and glucose in protoplast culture medium was beneficial to increase division frequency and plating efficiency. The calli were allowed to grow to 4--6 mm in height with red colour and compact structure on the gelrite-sohdified NLZ1 proliferation medium in 3 weeks and were transferred onto NLF differentiation medium where the frequency of shoot formation could reach 100%. The 3 cm high shoots were then cut off from the callus and rooted on 1/2 MS medium.
Abstract (Browse 1812)  |  Full Text PDF       
Effect of ABA on the in Vitro Induction of Floral Buds of Dendrobium candidu Wall.ex Lindl.
Author: Wang Guang-yuan, Liu Pei, Xu Zhi-hong and Cai Nan-hai(Chua Nam-hai)
Journal of Integrative Plant Biology 1995 37(5)
    An in vitro flowering system of Dendrobium candidurn Wall. ex Lindl., a wild species of orchid, was established. Callus was induced from seeds and protocorms formed on MS agar medium supplemented with 0.3 mg/L NAA under diffused light. Floral buds were induced when protocorms were cultured on MS agar medium supplemented with 2 mg/L 6-BA and 0.5 mg/L NAA, the frequency of floral bud induction being 27.0%. When protocorms were precultured on MS medium supplemented with 0. 5 mg/L ABA for 15 days and then transfered onto MS medium supplemented with 2 mg/L 6-BA, the frequency of floral bud induction increased greatly reach 84.0 % during a period of 5 months. Meanwhile the number of branches and floral buds also increased and in some instances inflorescence appeared. Never the less, no floral bud was formed if protocorms continued to be cultured on the ABA-containing MS medium.
Abstract (Browse 1943)  |  Full Text PDF       
Effect of ABA on the Activity of Mitochondrial Membrane Bound Na+-K+ ATPase
Author: Chen Kao-shan and Zhou Xie
Journal of Integrative Plant Biology 1995 37(5)
    Effect of ABA on the activity of mitochondrial membrane bound Na+-K+ATPase during isolation of mitochondria from soybean cotyledons, there was an increased activity of the mitochondrial membrane bound Na+-K+ATPase if abscisic acid (ABA) was added to the medium when soybean seedling were grown at 27 or 16, 40 mol/L ABA could change the turning point temperature of Arrhenius the activation energies (Ea) of Na+-K+ATPase from 36.6 or 22.7 decreased to 30. 3 or 17.8 respectively. The Km value and S0.5 value for this enzyme with ABA was higher than that without ABA. Hill coefficient (n) of this enzyme with ABA was 1.01 and without ABA was 1.89. The o2 uptake of mitochondria also increased. These results showed that the temperature of phase transition of mitochondrial membrane were decreased by ABA treatment.
Abstract (Browse 1778)  |  Full Text PDF       
Study of Glutamyl Kinase in Rice
Author: He Dao-yao
Journal of Integrative Plant Biology 1995 37(5)
    Proline synthesis and accumulation has often been shown to occur in plants as a consequence of environmental stress. The physiological significance of proline accumulation is assumed to be associated with the suggested ability of proline to act as an osmoregulator, protective agent or storage compound. Despite the importance of this compound, however, the exact metabolic route and enzymes involved in the synthesis of proline in plants have not been unequivocally identified. The proline synthesis route in plants is thought to resemble the pathway in bacteria. With the exception of 1-pyrroline-5-carboxylate reductase (P5CR), which has been recently characterized, little is known about the other two enzymes, glutamyl kinase (GK) and -glutamyl phosphate reductase (GPR). This present paper provides the preliminary results of GK activity assay and its partial purification in rice (Oryza sativa L. ). The enzyme activity assayed was quite low. Different saturation of (NH4)2SO4 was used for precipitation of sample proteins, the enzyme was enriched in the fraction of 30%--60% (NH4)2SO4 saturation and showed a two-fold increase in the assayed activity. The enzyme activity was inhibited by the addition of proline from 10 to 1000 mmol/L. The result indicated that the enzyme from rice was less sensitive to feedback inhibition by proline than that in bacteria. The enzyme was primarily purified by Sephadex G- 150 chromatography. Two activity peaks were discovered in the fractions.
Abstract (Browse 1777)  |  Full Text PDF       
Pollination-Induced Expression of Ethylene Biosynthetic Genes at Transcription Level in Orchid Flowers
Author: Zhang Xian-sheng, Yan Xian-xi and Ma Xiao-jie
Journal of Integrative Plant Biology 1995 37(5)
    The authors investigated pollination-induced ethylene production and expression patterns of genes encoding 1-aminocyclopropane-l-carboxylate (ACC) synthase and ACC oxidase in orchid flowers (Doritaenopsis hybrida Hort. ). Following pollination both ACC synthase and ACC oxidase mRNAs were detected in the different organs of flowers, and the patterns of both ACC synthase and ACC oxidase mRNA accumulation were similar, mRNA accumulation of ACC synthase mRNA was more organ-specific than that of ACC oxidase mRNA. However, ACC oxidase mRNAs were much more abundant than ACC synthase mRNAs in the flower organs.
Abstract (Browse 1794)  |  Full Text PDF       
Genetic Analysis of Null Alleles in Rice Doubled Haploid Plants from Anther Culture
Author: Li Ping, Zhu Li-huang, Chen Ying, Zhou Kai-da and Lu Chao-fu
Journal of Integrative Plant Biology 1995 37(5)
    In the research of constructing a rice(Oryza sativa) molecular map, 4 RFLP markers, i.e. RG 229, RG 419, RG 424 and RG 353, detected the null alleles in the indica and japonica parental rice. RG 229 indicated two null alleles in indica rice Gui 630, and each of the other markers revealed a null allele in japonica rice 02428. Genetic analysis in the doubled haploid (DH) population consisting of 81 plants showed that the linkage relationships between these null allele loci and the neighboring molecular markers shown in McCouch' s rice molecular map were changed. In addition, the marker RG 684 could detect its null alleles in some DH plants, though the RG 684 sequence did exist in the genomes of both parents. Appearence of null alleles might be induced by transpositional changes on chromosomes.
Abstract (Browse 1933)  |  Full Text PDF       
Light-Induced Damage of Pheophytin a Molecule in the Isolated Photosystem Reaction Center D1/D2/Cyt b559 Complex
Author: Kuang Ting-yun, Hou Jian-min, Peng De-chuan, Tang Chong-qin and Tang Pei-song
Journal of Integrative Plant Biology 1995 37(5)
    Photodamage of some pigments in the isolated photosystem (PS ) reaction center D1/D2/Cyt b559 complex from spinach has been investigated by means of high performance liquid chromatography. The light-induced damage of pheophytin a (pheo a) in the complex was observed for the first time. The content of pheo a decreased about 47 % by illumination, suggesting only one of the two pheo a molecules in the PS reaction center complex was damaged. No damage of -carotene was found.
Abstract (Browse 1685)  |  Full Text PDF       
Preliminary Study of the Benthic Marine Algae on the Northern Bohai Sea Coast
Author: Sui Zhan-ying and Fu Jie
Journal of Integrative Plant Biology 1995 37(5)
    The composition, distribution and seasonal variation of the benthic marine algae in the northern Bohai Sea coast were studied. 119 species of benthic marine algae were found in the northern Bohai Sea coast. They belong respectively to Cyanophyta, Thodophyta, Phaeophyta and Chlorophyta. The study showed that the marine algae flora of the northern Bohai Sea coast belongs to warm temperate in nature with considerable subtropical composition. There were 57 species of economic algae in the northern Bohai Sea coast and 7 newly recorded species.
Abstract (Browse 1673)  |  Full Text PDF       


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