Effects of Supplementation of Various Medium Components on Chinese Hamster Ovary Cell Cultures Producing Recombinant Antibody

Thirteen vitamins, twenty amino acids, hormones, inorganic salts, and other chemical agents, which constitute typical serum-free media, were evaluated for the development of fortified medium to enhance cell growth and productivity of recombinant antibody in the cultures of the recombinant Chinese hamster ovary (rCHO) cells. Two different rCHO cell lines, rCHO-A producing recombinant antibodies against the human platelet and rCHO-B secreting recombinant antibodies against the S surface antigen of Hepatitis B, respectively, were cultivated in batch suspension mode. Concentration of interested component in the tested medium was doubled to examine the fortification effect. Growth of rCHO-A cell and its antibody production were slightly improved with addition of either choline chloride, folic acid, thiamine⋅HCl, or Long^TMR³IGF-I. On the other hand, in the cultivation of rCHO-B cell which was more sensitive to its environmental changes, hormones such as Long^TMR³IGF-I and triiodothyronine (T₃) as well as various vitamins involving choline chloride, i-inositol, niacinamide, pyridoxine HCl, and thiamine⋅HCl enhanced the cell growth and antibody production. Particularly, when concentration of consuming amino acid was doubled, remarkable increase in specific productivity was served, resulting in high final antibody concentration. These results were believed to provide a fundamental strategy of medium fortification useful for improvement of recombinant antibody production in serum-free medium. These authors contributed equally to this work     

Effect of Various Elicitors on Lignan Biosynthesis in Callus Cultures of Linum austriacum

Effects of elicitors (mannan, -1,3-glucan, and ancymidol) on the activity of several key enzymes participating in lignan biosynthesis were studied in Linum austriacum L. cell cultures. The activities of L-phenylalanine ammonia-lyase, polyphenoloxydase, tyrosine ammonia-lyase, soluble phenoloxidase, and membrane-bound and soluble oxidases were assayed. The elicitors under study affected various steps in the metabolic pathway of lignan biosynthesis. Elevated enzyme activity accompanied an elicitor-enhanced synthesis of podophyllotoxins and peltatins.     

Effects of Various Elicitors on the Transcription of a β-1,3-endoglucanase Gene in Citrus Fruit

Very little is yet known regarding the molecular mechanisms involved in pathogen defense responses in citrus fruit. Recently, a basic β-1,3-endoglucanase (EC 3.2.2.39) belonging to the pathogenesis-related (PR) group of proteins, has been purified from Citrus sinensis (L) Osbeck cv. `Valencia' orange callus. Specific antibodies raised against the purified protein were used to screen `Valencia' callus and flavedo cDNA expression libraries, and to isolate its corresponding cDNA, designated gns1. The gns1 gene encodes a predicted polypeptide of 336 amino acids with a molecular mass of 37.3 kDa and a basic pI of 9.19, and shares 55–65% identity with several other plant β-1,3-endoglucanase proteins. Hereby, we show that the expression of the gns1 gene is markedly induced by wounding and inoculation with Penicillium digitatum (Pers. Fr.) Sacc., and following treatments with various elicitors that induce fruit resistance against P. digitatum . These treatments include UV irradiation, application of jasmonic acid (JA), β-aminobutyric acid (BABA), Candida oleophila antagonist yeast cells and hot water rinsing and brushing. Overall, based on various RNA gel blot hybridizations, we assume that gns1 is most likely to be part of the molecular mechanisms involved in pathogen defense responses in citrus fruit. *     

In vitro Propagation of Ginkgo biloba by Using Various Bud Cultures

The purpose of this research is micropropagation of Ginkgo biloba L. Apical and nodal meristems removed from plantlets or apical buds from a tree were used as explants. Meristems produced an extensive callus and single or rare multiple shoots on Murashige and Skoog medium with different growth regulators and endosperm extract (En) obtained from mature seeds of the same species. For successful root production it was necessary to transfer the shoots to a rooting medium with En. This revised version was published online in July 2006 with corrections to the Cover Date.     

Mineral Content of the Pulp and Peel of Various Citrus Fruit Cultivars

Biological Trace Element Research - The aim of the study was to compare the mineral content between the peel and the pulp of citrus fruits and to determine which citrus fruit, among orange (Citrus...     

Toxic and Protective Effects of l-DOPA on Mesencephalic Cell Cultures

Abstract: The autoxidation of L-DOPA or dopamine (DA) and the metabolism of DA by monoamine oxidase generate a spectrum of toxic species, namely, hydrogen peroxide, oxy radicals, semiquinones, and quinones. When primary dissociated cultures of rat mesencephalon were incubated with L-DOPA (200 μ M ) for 48 h, the number of tyrosine hydroxylase-positive neurons (DA neurons) was reduced to 69.7% of control values, accompanied by a decrease in [³H]DA uptake to 42.3% of control values; the remaining DA neurons exhibited reduced neurite length and overall deterioration. Lack of simultaneous change in the number of neurons stained with neuron-specific enolase indicated that toxicity was relatively specific for DA neurons. At the same time, the level of GSH, a major cellular antioxidant, rose to 125.2% of control values. Thus, exposure of mesencephalic cultures to L-DOPA results in both damaging and antioxidant actions. Ascorbate (200 μ M ), an antioxidant, prevented the rise in GSH. The effect of ascorbate on GSH points to an oxidative signal to initiate the rise in GSH content. On the other hand, neither inhibition of monoamine oxidase with pargyline nor addition of superoxide dismutase or catalase to the culture medium prevented the rise in GSH level or the loss in [³H]DA uptake. The latter results tend to exclude the products of monoamine oxidase activity or the presence of hydrogen peroxide or superoxide in the medium as responsible agents for the rise in GSH or neuronal toxicity. In cultures treated with L-buthionine sulfoximine (L-BSO), an inhibitor of GSH synthesis, l-DOPA prevented cell death by L-BSO.     

Volatile Compounds Originating from Mixed Microbial Cultures on Building Materials under Various Humidity Conditions

We examined growth of mixed microbial cultures (13 fungal species and one actinomycete species) and production of volatile compounds (VOCs) in typical building materials in outside walls, separating walls, and bathroom floors at various relative humidities (RHs) of air. Air samples from incubation chambers were adsorbed on Tenax TA and dinitrophenylhydrazine cartridges and were analyzed by thermal desorption-gas chromatography and high-performance liquid chromatography, respectively. Metabolic activity was measured by determining CO₂ production, and microbial concentrations were determined by a dilution plate method. At 80 to 82% RH, CO₂ production did not indicate that microbial activity occurred, and only 10% of the spores germinated, while slight increases in the concentrations of some VOCs were detected. All of the parameters showed that microbial activity occurred at 90 to 99% RH. The microbiological analyses revealed weak microbial growth even under drying conditions (32 to 33% RH). The main VOCs produced on the building materials studied were 3-methyl-1-butanol, 1-pentanol, 1-hexanol, and 1-octen-3-ol. In some cases fungal growth decreased aldehyde emissions. We found that various VOCs accompany microbial activity but that no single VOC is a reliable indicator of biocontamination in building materials.     

The Effect of Various Mono–and Disaccharides on the Growth of Sphagnum nemoreum Thalli in Sterile Cultures

Sphagnum nemoreum Scop. thalli were grown under sterile conditions in order to study their ability to use certain carbohydrates for their growth, with special reference to the sugars occurring free in Sphagnum peat. It was found that Sphagnum nemoreum thallus could not grow at ail in vitro unless some organic carbon source war present. Growth was dependent on both the quality and the quantity of the sugar. Sucrose (1 per cent) proved to be the best carbon source and light intensity did not have any marked influence on growth under these conditions. Glucose and fructose were also growth promoting. Glucose is the main free sugar in the peat. Mannose (0.25 per cent) was almost as good a carbon source as sucrose in the same concentration. It is known that mannose accumulates in the Sphagnum peat during humification. and so does rhanmose, which was utilized to some extent. These two sugars are inhibitory to many mono–and dicotyledons. Ribose, galactose. arabinose and xylose were found to be toxic to Sphagnum.     

Differences in Strain Virulence of the European Stone Fruit Yellows Phytoplasma and Susceptibility of Stone Fruit Trees on Various Rootstocks to this Pathogen

Twenty strains of the European stone fruit yellows (ESFY) phytoplasma showed great differences in virulence when examined by graft inoculation of trees on peach, peach hybrid GF 677 and P. 'Marianna' GF 8/1 rootstocks. The most virulent strains killed all trees on peach rootstocks whereas the mild strains did not cause mortality but induced only mild foliar symptoms and slightly reduced vigour. Virulence often depended on the pathogen–scion combination and was in several cases most severe when the scion consisted of the original host of the pathogen. To examine resistance in stone fruits, trees on a total of 23 rootstocks were inoculated with the ESFY strains. Trees on the Prunus domestica stocks Ackermann's, Brompton and P 1275 and on Prunus cerasifera stock Myrabi were little affected. Slightly more damage occurred in trees on rootstocks GF 677, GF 8–1, and the P. insititia stocks St Julien A and St Julien GF 655/2. Ishtara, P. cerasifera stock Myrobalan, and peach rootstocks Higama and GF 305 were shown to be moderately susceptible and a high susceptibility was found in trees on peach rootstocks Montclar, peach seedling, Rutgers Red Leaf, and Rubira, on apricot seedlings and St Julien 2. Of flowering cherry trees on various rootstocks, the least susceptible were those on Gisela 3 and F 12/1 whereas Gisela 1, Weihroot 158 and Gisela 5 were more affected. Phytoplasmas were detected by either DAPI (4'-6-diamidino-2-phenylindole) staining or polymerase chain reaction in all rootstocks and scions tested. However, detection frequency and phytoplasma concentrations were usually lower in the more tolerant hosts than in susceptible genotypes.     

Effects of Activated Charcoal on Growth and Morphogenesis in Cell Cultures

The effects of activated charcoal on growth and morphogenesis in plate cultures of different plant cells have been studied. It was shown that medium containing charcoal induced embryogenesis in cultures of Daucus carota in which embryo formation could not be brought about by omitting auxin from the medium. Charcoal-medium also induced abundant root formation in older cultures of Allium cepa, which normally did not produce roots. The growth of cultures of Glycine max and Haplopappus gracilis was totally inhibited by charcoal. It is thought that activated charcoal removes substances from the medium, one of which might be auxin.     

不同纬度产地纽荷尔脐橙果实品质状况初步研究

以纽荷尔脐橙为试材,分析了我国甜橙优势区域内不同纬度地区纽荷尔脐橙果实主要的品质指标。结果表明,纽荷尔脐橙果实的多数品质指标随纬度的变化呈现有规律的变化;线性回归分析结果显示,纽荷尔脐橙果实几项品质指标与纬度的相关性为:可溶性固形物含量呈显著线性正相关,可滴定酸含量呈极显著线性正相关,固酸比呈极显著线性负相关,果皮黄色度和着色强度呈极显著线性负相关,色调角呈显著线性负相关,VitC含量与纬度的线性关系接近显著程度。     

Nicotine Production by Tissue Cultures of Tobacco as Influenced by Various Culture Parameters

Assimilate efflux from vacuum-infiltrated leaf slices (spinach, barley) into a buffered solution was examined in relation to Ca^{+ +} -activity and osmotic conditions. Efflux from isolated mesophyll protoplasts and from a unicellular green alga (Eremosphaera viridis de Bary) was also measured.In the presence of Ca^{+ +}, assimilate efflux from leaf slices was small (1 to 5 % of the total carbon fixation rate, depending on osmotic conditions). Efflux was drastically stimulated by addition of Ca^{+ +} -chelators. If expressed as µmol carbon mg^-1 chlorophyll h^-1, it reached 50 % of the assimilation rate. Efflux from protoplasts or algae was slow and insensitive to Ca^{+ +} chelators at concentrations which caused fast efflux from leaf slices.Assimilate efflux from leaf slices was rather unspecific. Both in the tissue and the surrounding medium, sucrose was the most abundantly labelled compound (70 to 80 % of total soluble labelled material).A 50 % decrease of efflux was observed when turgor pressure was lowered by addition of sorbitol (200 to 300 mosmol kg^-1). At extremely high sorbitol concentrations (> 1500 mosmol kg^-1) efflux increased again and was relatively less stimulated by EDTA.It is suggested that assimilate efflux from leaf slices is mainly diffusion through open veins and/or plasmodesmata. When these symplastic connections are closed by addition of Ca^{+ +}, the remaining transmembrane flux into the apoplast is small. Thus, assimilate movement from the mesophyll to the phloem appears to be symplastic, not apoplastic as suggested in the literature.     

植物生长调节剂对火龙果坐果率和果实生长的影响

以‘红龙果Ⅱ’为试材,研究不同生长调节剂处理对火龙果坐果率和果实生长的影响。结果表明：浓度为30 mg/L GA3或100 mg/L 6-BA对提高火龙果坐果率及促进果实生长效果最佳。     

Effective Trapping of Fruit Flies with Cultures of Metabolically Modified Acetic Acid Bacteria

Acetoin in vinegar is an attractant to fruit flies when combined with acetic acid. To make vinegar more effective in attracting fruit flies with increased acetoin production, Komagataeibacter europaeus KGMA0119 was modified by specific gene disruption of the acetohydroxyacid isomeroreductase gene (ilvC). A previously constructed mutant lacking the putative ligand-sensing region in the leucine-responsive regulatory protein (KeLrp, encoded by Kelrp) was also used. The ilvC and Kelrp disruptants (KGMA5511 and KGMA7203, respectively) produced greater amounts of acetoin (KGMA5511, 0.11%; KGMA7203, 0.13%) than the wild-type strain KGMA0119 (0.069%). KGMA7203 produced a trace amount of isobutyric acid (0.007%), but the other strains did not. These strains produced approximately equal amounts of acetic acid (0.7%). The efficiency of fruit fly attraction was investigated with cultured Drosophila melanogaster. D. melanogaster flies (approximately 1,500) were released inside a cage (2.5 m by 2.5 m by 1.5 m) and were trapped with a device containing vinegar and a sticky sheet. The flies trapped on the sticky sheet were counted. The cell-free supernatant from KGMA7203 culture captured significantly more flies (19.36 to 36.96% of released flies) than did KGMA0119 (3.25 to 11.40%) and KGMA5511 (6.87 to 21.50%) cultures. Contrastingly, a 0.7% acetic acid solution containing acetoin (0.13%) and isobutyric acid (0.007%), which mimicked the KGMA7203 supernatant, captured significantly fewer flies (0.88 to 4.57%). Furthermore, the KGMA0119 supernatant with additional acetoin (0.13%) and isobutyric acid (0.007%) captured slightly more flies than the original KGMA0119 supernatant but fewer than the KGMA7203 supernatant, suggesting that the synergistic effects of acetic acid, acetoin, isobutyric acid, and unidentified metabolites achieved the efficient fly trapping of the KGMA7203 supernatant.     

注射施肥对桃园土壤环境和果实产量品质的影响

以盛果期10a生品种‘阿部白’桃为试材,在大田条件下,进行连续2年定位施肥试验,设置放射沟施全量肥(R_(NPK))、放射沟施半量肥(R_(1/2NPK))、注射施全量肥(I_(NPK))和注射施半量肥(I_(1/2NPK))4个处理,分析注射施肥对桃园0~60cm土层土壤养分含量、土壤酶活性和微生物数量以及果实品质的影响,探讨注射施肥在桃树生产中的应用,为高效施肥和提高水肥利用率提供理论依据。结果表明:(1)桃幼果膨大期(5月15日),0~40cm土层中,试验各处理的总氮钾含量差异不明显,注射施肥土壤中速效养分含量显著低于放射沟施肥;果实成熟前期(7月15日),在相同施肥量下,0~40cm土层中注射施肥处理的碱解氮、速效钾含量均以I_(NPK)处理的最高,且分别比放射沟施肥处理显著提高16.61%、12.56%和10.15%、23.93%,同时注射施肥处理的0~40cm土层中有机质含量均低于放射沟施肥处理,部分处理差异达显著性水平;不同处理对土壤中磷的影响较小。(2)果实成熟前期,0~60cm土层中注射施肥土壤中蔗糖酶、过氧化氢酶和脲酶活性,以及0~20cm土层的碱性磷酸酶均显著高于放射沟施肥;同期根际土壤微生物数量变化规律与土壤酶活性变化基本一致。(3)INPK处理的桃单果重、可溶性固形物含量显著高于其他施肥处理,且分别比R_(NPK)处理显著增加了12.24%和11.21%;I_(1/2NPK)处理的单株果实产量最高(65.25kg)并显著高于放射沟施肥处理,但与INPK处理间差异不显著。研究认为,注射施肥对桃树土壤中养分含量、土壤酶活性和微生物数量以及果实品质都有积极的影响,且以NPK全量注射施肥处理效果最佳。     

Aminoethoxyvinylglycine Effects on Late-Season Apple Fruit Maturation

Aminoethoxyvinylglycine (AVG) inhibits 1-aminocyclopropane-1-carboxylic acid (ACC) synthase, and thus blocks ethylene synthesis. Preharvest foliar application of AVG to apple (Malus domestica Borkh.) fruit retards several key events of maturation including climacteric ethylene production, starch conversion to sugars, fruit softening, and abscission zone development. Although the impact of AVG on apple fruit maturation is well known, the biochemical basis of these effects is not well understood. The effects of AVG application on Redchief Delicious apple fruit maturation were studied. AVG applied four weeks prior to harvest significantly reduced internal ethylene levels, amylose degradation, and accumulation of sucrose, glucose, and sorbitol. Because AVG application coincidentally inhibited starch degradation and the increase in internal ethylene, we investigated the enzymatic basis of starch mobilization in apple fruit. Amylase activity was somewhat reduced in AVG-treated fruit. Amylase activity was less in AVG-treated fruit during the early stages of starch mobilization. Starch phosphorylase activity increased dramatically during the later stages of starch mobilization, but was not affected by AVG treatment. Soluble starch synthase activity was also unaffected by AVG treatment and remained constant throughout the eight-week harvest period. Moreover, AVG did not affect the levels of amylopectin, fructose, malate, ascorbate, citrate, or anthocyanin. These results suggest that apple fruit ripening has both ethylene-dependent and -independent processes occurring simultaneously.     

Comparison of Various Methods for Preparation of Viral Serological Antigens from Infected Cell Cultures

In efforts to prepare more potent and sensitive viral serological antigens, several aspects of the production of antigens from infected cell cultures were studied. Antigens derived from whole, infected culture material and from the cellular and fluid phases were compared. Freezing and thawing, sonication, and alkaline buffer extraction were compared for effectiveness in releasing antigen from host cells. The effect of the multiplicity of infection on titers of viral antigens produced in cell cultures was studied. Generally, higher titered antigens were derived from the infected cells than from the culture fluids, but for certain viruses complement-fixing (CF) antigens derived from the culture fluids gave higher antibody titers than did cell-associated antigens. With each virus-host cell system studied, treatment with alkaline buffers extracted appreciable amounts of CF antigen from the host cells, but in some instances more antigen was released by freezing and thawing or by sonication. Extraction of infected cells with alkaline buffers was not a satisfactory method for preparation of hemagglutinating (HA) antigens for any of the viruses studied. The highest-titered HA antigens were produced from infected cells disrupted by freezing and thawing or sonication. The highest titered CF and HA antigens were produced from cell cultures infected at multiplicities of one or greater. Complement-fixing antigens produced by infecting cells in suspension and then planting had lower titers than antigens produced in parallel by infecting developed monolayers. Optimal methods are summarized for preparation of serological antigens to a variety of viruses of man.     

Effects of Sulphur-containing Compounds on Lettuce Fruit Germination

The germination of Lactuca sativa L. cv. Arctic King fruit ona 0.5 per cent agar substrate was inhibited in light at temperaturesabove 31?C (? 1.0?). This precisely repeatable value, termedthe upper temperature cut-off point, is characteristic for thislettuce variety. The position of this cut-off point is raisedby the presence of thiourea (500–1000 mg/1) and a numberof other thiol compounds. This effect is not shown by all thiolcompounds and is dependent on the configuration of the restof the molecule. The presence of a hydroxyl group in the testcompound often results in the temperature cut-off point beinglowered but again this effect depends on the configuration ofthe rest of the molecule. It is postulated that the stimulatingeffects of active thiol compounds are related to hydrophobicinteractions with a macro-molecule involved in the initiationof germination.     

Computerized Optimization of the Relative Growth of Callus Cultures of Orange Fruit Tissues7

The induction and growth of callus cultures from different partsof the fruit of Coorg mandarin orange (Citrus reticulata, Blanco)and the hormonal combinations that can maximize the growth witha basal MS medium, have been explored using Response SurfaceMethodology. The relative growth, protein, peroxidase pattern,and GC fingerprinting of volatiles have been studied. Calluscultures from juice vesicles were found to be more similar totheir explant counterpart than those from albedo, for all propertiesstudied. Key words: Callus culture, Citrus, albedo, juice vesicles, response surface methodology, growth optimization