Co-immobilization of dextransucrase and dextranase in alginate

Dextransucrase from Leuconostoc mesenteroides and dextranase from Penicillium lilacinum were co-immobilized and used to produce isomaltooligosaccharides from sucrose. The enzymes were co-immobilized by encapsulating soluble dextransucrase and dextranase covalently attached to Eupergit C in alginate (beads, fibers, and capsules). The alginate capsule co-immobilization was done in the presence of soluble starch and resulted in a high immobilization yield (71%), and the enzymes retained their activities during 20 repeated batch reactions and for a month in storage at 4 °C. The presence of starch was essential for the stability of dextransucrase in alginate capsules. Furthermore, it is important that the dextranase be pre-immobilized prior to alginate capsule co-immobilization to prevent dextranase leakage and inactivation of dextransucrase. The co-immobilized enzymes formed oligosaccharides from sucrose, which can be used as prebiotics. In addition, the oligosaccharides that were produced after the addition of sucrose reacted with the alginate fiber-encapsulted dextransucrase, thus increasing the amount of prebiotics. Co-immobilization in alginate fiber and beads also resulted in high yields (70 and 64%), but enzymatic activities decreased by 74 and 99%, respectively, after a month in storage at 4 °C. The newly developed alginate capsule method for co-immobilization of dextransucrase and dextranase is simple yet effective and has the potential for industrial-scale production of isomaltooligosaccharides.     

粉被虫草静置发酵产N6-(2-羟乙基)腺苷培养基组分及前体物的筛选

在静置培养条件下,对粉被虫草cp菌株产N6-(2-羟乙基)腺苷（简称HEA）的培养基及其组分进行优化。采用单因素实验方法,以HPLC法对cp菌丝体中HEA进行检测,以HEA的产量为指标,结果表明查氏培养基有利于cp菌株产HEA,其产量是沙氏和PD培养基的3.7倍和4.48倍。以查氏培养基为基础培养基,进行碳、氮源的筛选中,最有利于cp菌株产HEA的是蔗糖和硝酸钠;在无机盐的筛选中,K2HPO4是促进cp菌株累积HEA最主要的无机盐,其HEA产量较CK增长了5 822.37%（提高了58.2倍）;在查氏培养基上添加不同前体物和氨基酸,发现其结构类似物腺苷、次黄嘌呤和腺嘌呤都有促进cp菌株累积HEA的能力,其中腺苷和次黄嘌呤能提高其产量60%以上;而组氨酸是最有利于cp菌株累积HEA的氨基酸,产量达到（45.56±2.8）mg/L,较CK提高HEA产量251.68%,其次是L-谷氨酸增产184.19%。     

A STUDY OF A GRAM-POSITIVE COCCUS RESPONSIBLE FOR ROPINESS AND VISCOSITY IN MALT WORT

SUMMARY: A Gram-positive coccus with the properties typical of pediococci produced a ropy, viscous growth in malt wort. Nutritional studies in the media of Dunn et al. (1947) showed pantothenic acid to be required by the organism but growth occurred in the complete medium only when a trace of malt wort was present. Seven other vitamins tested were not necessary for growth.
In a medium consisting of mineral salts, glucose and alcoholic extracts of peptone and yeast autolysate the organism grew well and the viscosity of the medium increased. Higher viscosities were attained when the buffering capacity of the medium was increased either by raising the concentrations of the alcoholic extracts or by adding sodium acetate. The material responsible for the viscosity was produced in small amounts only and appeared to be a mucopolysaccharide. The only sugar obtained on hydrolysis of this material was glucose.
Fructose, maltose, sucrose and raffinose were all fermented by the organism but only in media containing maltose was the increase in viscosity comparable with that attained in a glucose-containing medium.     

Culture-induced changes in osmolality of tobacco cell suspensions using four exogenous sugars

Suspension cultures of tobacco cells were grown in B5 media supplemented with sucrose, glucose, mannitol and sorbitol as exogenous sugars to examine culture-induced changes in the osmolality of the medium. Osmolality decreases were greatest in sucrose and glucose media during the 14 days in culture, and in glucose media were essentially linear, presumably reflecting the use of this sugar as a food source. Osmolality decreases occurred during the first week of culture in mannitol- and sorbitol-supplemented media, but later stabilized. Fresh weight of cultured cells in sucrose- and glucose-supplemented media increased by <200% during 14 days in culture, whereas cultured cells in mannitol- and sorbitol-supplemented media increased by only 39 and 48%, respectively. Cells transferred to the original liquid medium (B5 medium with 3% sucrose and 3% glucose) grew vigorously if they had been cultured in sucrose- and glucose-supplemented media; however, cells grown in mannitol- and sorbitol-supplemented media needed to be subcultured several times to recover their normal growth rate. By subculturing cells into increasingly higher conditions of sugars, cells tolerant to 560 mOsmol kg-1 H2O were obtained. The high osmolality-adapted cells increased by 140% in fresh weight in 8% glucose-supplemented medium. Glucose was best suited for producing the high osmolality required because sucrose concentrations at 10% sucrose and above resulted in cell death. To limit the decrease of osmolality in these suspension cultures requires changing the medium every 3 days to maintain osmolality above the 530 mOsmol kg-1 H2O needed to co-culture these as feeder cells with gametic and zygotic cells. This revised version was published online in June 2006 with corrections to the Cover Date.     

海滨锦葵胚轴愈伤组织诱导及植株再生

以海滨锦葵(Kosteletzkya virginica)胚轴为外植体, 在9种不同激素配比的培养基上进行愈伤组织诱导、继代培养、不定芽分化及生根培养, 确定了植株再生的最适培养条件: (1)愈伤组织诱导最适培养基为MS + IAA 1.0 mg.L-1 + KT 0.3 mg.L-1 + sucrose 30 g.L-1 + agar 8 g.L-1, 愈伤组织诱导率为93.94%; (2)不定芽诱导最适培养基为MS + IAA 0.1 mg.L-1 + ZT 0.5 mg.L-1 + sucrose 30 g.L-1 + agar 8 g.L-1, 不定芽诱导率为65.83%; (3)生根最适培养基为MS + sucrose 30 g.L-1 +agar 8 g.L-1, 生根率为96.67%。炼苗移栽后, 成活率可达85%。     

淡紫拟青霉右旋糖酐酶的形成条件

比较了各种碳水化合物对淡紫拟青霉(Paecilomyces lilacinus)右旋糖酐酶形成的影响,右旋糖酐是最好的碳源,也是最佳诱导物。不同分子量(17．2—1000kD)的右旋糖酐对酶形成的诱导作用不同,酶的产生随右旋糖酐分子量的增大而增加。用分子量为1000kD的右旋糖酐作碳源时比用17.2kD的右旋糖酐作碳源时的产酶量高40％以上。用右旋糖酐和其它糖的混合物作碳源时,酶的形成受到不同程度的抑制。右旋糖酐酶形成的其它适宜条件：氮源为牛肉蛋白胨,培养基初始pH6．0—7.0.种龄为48小时,在250ml三角瓶中装50ml培养基,于28℃在200r／min摇床上培养6天。     

百脉根愈伤组织原生质体再生植株

百脉根无菌苗幼茎在含２．０ｍｇ／Ｌ－,２,４－Ｄ,０．１ｍｇ／Ｌ２－ｉｐ的ＭＳ培养基上诱导和继代培养愈伤组织。选取绿色松散颗粒愈伤组织分离原生质体。原生质体培养在调整珠ＫＭ８Ｐ,Ｖ－ＫＭ,ＭＳ和ＳＨ培养基上「含３００ｍｇ／Ｌ,ＣＨ,２％ＣＷ,２％蔗糖,６％葡萄糖,２．０ｍｇ／Ｌ,２,４－Ｄ,０．５ｍｇｇ／Ｌ,ＢＡ,５ｍｍｏｌ／Ｌ ＭＥＳ」,原生质体再生细胞均能分裂,并形成小愈伤组织,但以ＫＭ８０为     

Studies of protoplast culture types and plant regeneration from callusderived protoplasts of Asparagus officinalis L. cv. Lucullus 234

Plating efficiency and colony formation of callus-derived protoplasts of Asparagus officinalis L. cv. Lucullus 234 differed significantly with different protoplast culture media and types of culture. Osmotic conditions and hormone concentrations of liquid media produced the greatest influence on plating efficiency and colony formation in bead culture. Protoplasts grew best in bead culture with a solid modified Kao & Michayluk protoplast culture medium (KM) supplemented with 0.5 mg l^–1 -naphthaleneacetic acid (NAA), 0.5 mg l^–1 2,4-dichlorophenoxyacetic acid (2,4-D), 0.5 mg l^–1 kinetin, and 0.6% agarose (KM6) and a liquid modified KM medium differing from KM6 medium in sugar content, having 0.18 M sucrose and 0.18 M mannitol (A8). An average plating efficiency of 19.1% and colony formation of 15.5% was obtained one week after isolation in bead culture with the KM6 and A8 media. The highest average shoot regeneration of 92.3% was obtained with a Murashige & Skoog medium (MS) containing 0.125 mg l^–1 NAA, 0.125 mg l^–1 2,4-D, 0.25 mg l^–1 6-benzylaminopurine (6-BAP) and 3% sucrose. Plants have been regenerated and transferred to the greenhouse.Abbreviations NAA -naphthaleneacetic acid - 2,4-D 2,4-dichlorophenoxyacetic acid - 6-BAP 6-benzylaminopurine     

Glucosyltransferase Mutants of Leuconostoc mesenteroides NRRL B-1355

Leuconostoc mesenteroides NRRL B-1355 produces dextrans and alternan from sucrose. Alternan is an unusual dextran-like polymer containing alternating α(1→6)/α(1→3) glucosidic bonds. Cultures were mutagenized with UV and ethyl methanesulfonate, and colony morphology mutants were selected on 10% sucrose plates. Colony morphology variants exhibited changes from parent cultures in the production of one or more glucosyltransferases (GTFs) and glucans. Mutants were characterized by measuring resistance of glucan products to dextranase digestion, by electrophoresis, and by high-pressure liquid chromatography of maltose acceptor products generated from sucrose-maltose mixtures. Some mutants produced almost pure fraction L dextran, and cultures exhibited a single principal GTF band on sodium dodecyl sulfate-acrylamide gels. Other mutants produced glucans enriched for alternan. Colony morphology characteristics (size, smoothness, and opacity) and liquid culture properties (clumpiness, color, and viscosity in 10% sucrose medium) were explained on the basis of GTF production. Three principal GTF bands were detected.     

Microviscosity and calcium exchange in yeast cells and effects of phenethyl alcohol

The microviscosity of the yeast cell interior determined by a fluorimetric method is 13.6 cp. Treatment with 0.5% phenethyl alcohol increases viscosity to 15 cp and increases calcium content fivefold. Phenethyl alcohol at low concentrations causes reversible increase in cell membrane permeability to fluorescein.     

Regulation of the activity of the alternative oxidase in callus forming discs from potato tubers

Callus-forming discs from potato tubers lose 80% of their starch during one month of incubation on nutrient medium containing either 0, 3 or 6% (w/v) sucrose. The content of soluble sugar in the discs varies from 5 mg (incubated without sucrose) to 22 mg (on 3% sucrose) and 40 mg (on 6% sucrose) per g fresh weight. The activity of the cytochrome pathway (V_cyt) increases during the first week of incubation on all media. Thereafter V_cyt decreases again on 0% sucrose medium, while it remains constant on 3 and 6% sucrose media. Alternative pathway capacity (V_alt), absent in freshly sliced tissue, shows a sharp increase during the first days of incubation, independent of the sucrose concentration in the medium. This capacity further increases during prolonged incubation on 3 and 6% sucrose but decreases on 0% sucrose. The in vivo activity of the alternative pathway (the participation in uninhibited respiration, ?V_alt) varies with the sucrose concentration and with the culture time. In tissue incubated for 2-3 weeks on 6% sucrose as much as 45% of the electrons are transfered to oxygen via the alternative pathway. In this tissue the factor Q (the part of the alternative pathway capacity that is operative) is about 0.8, while in tissue incubated on 0 and 3% sucrose media p generally does not exceed 0.5. When chloramphenicol, an inhibitor of mitochondrial protein synthesis, is added to the medium together with 3% sucrose, the increase in V_yet does not occur, while the induction of V_alt during the first week of incubation is the same as without chloramphenicol. A greater part of the alternative pathway capacity becomes operative in this tissue, leading to values of Q of almost 1 after prolonged incubation. Apparently, incubation on high sugar medium leads to extra participation in respiration of the energetically inefficient alternative oxidase pathway Excess sugar leads to wasteful respiration suggesting that the alternative oxidase functions as an ‘energy overflow’.     

Investigations into the Role of Sucrose in Potato cv. Estima Microtuber Production in vitro

Sucrose has been the carbohydrate traditionally used for potatomicrotuber production. Added to nutrient media, sucrose canact solely as a carbon source, or as an osmoticum, or both.Preliminary tests showed that the osmolarity of sucrose solutionswas increased by autoclaving, indicating some breakdown of thesugar. This was taken into consideration in experiments whichinvolved supplementing 4% sucrose media with sucrose, maltose,glucose or fructose, while keeping the osmotic potential ofthe media constant. A medium concentration of about 400 mM withonly sucrose was more suitable for microtuber production thanmedia supplemented with maltose, glucose or fructose. However,a better microtuber yield was obtained when hexoses were addedthan with unsupplemented 4% sucrose media. When glucose wassupplied at concentrations which had the same number of carbonatoms as 8% sucrose, the high osmolarity inhibited microtuberisation.Sugar movement in the tubering plantlet was followed using radio-labelledsucrose, glucose and fructose. The sucrose was translocatedand used at a faster rate than the other sugars, which tendedto remain in the roots of the plantlets. Furthermore, therewas no difference in microtuber production on media to whichthe sucrose was added before or after autoclaving, indicatingthat levels of breakdown were not severe enough to affect theprocess. Therefore, it is concluded that sucrose acts primarilyas a suitable carbon source for uptake and utilization by theplantlets, but, at 8%, it also provides a favourable osmolarityfor the development of microtubers.Copyright 1995, 1999 AcademicPress Solanum tuberosum (L.), potato, microtuber, media, sugar, sucrose, osmolarity, pH     

Stimulation of in vitro root and shoot growth of potato by increasing sucrose concentration in the presence of fluridone,an inhibitor of abscisic acid synthesis

Fluridone, an inhibitor of abscisic acid (ABA) biosynthesis, strongly stimulated rooting of nodal stem segments of potato (Solanum tuberosum L.) cultivar Arran Banner cultured in darkness on tuberisation medium. Inclusion of 10^-6 M ABA in the culture medium prevented this rooting response, indicating that root proliferation in the presence of fluridone could be due to inhibition of ABA synthesis. The rooting response to fluridone (increased total root number and root fresh weight) was obtained only at high sucrose concentrations (0.175 and 0.234 M) and was demonstrated with two potato cultivars and two culture media; one which favoured tuberisation and one which did not. Shoot numbers were also increased, but to a lesser extent than root numbers, and total fresh weight of plant material per culture was greatly increased by inclusion of both fluridone (10^-6 or 10^-5 M) and 0.234 M sucrose in the culture medium. The role of sucrose was not simply osmotic because when the osmolarity of fluridone medium was increased using mixtures of mannitol and sucrose, no root proliferation occurred unless sucrose predominated in the mixture.     

Sucrose-mediated giant cell formation in the genus Neisseria.

Growth of Neisseria perflava, Neisseria cinerea, and Neisseria sicca strain Kirkland in media supplemented with sucrose (0.5 to 5.0% w/v) resulted in the formation of giant cells. Response to sucrose was specific in that a variety of other carbohydrates did not mediate giant cell formation. Giant cells appeared only under growth conditions and did not lyse upon transfer to medium lacking sucrose or upon resuspension in hypotonic media. Reversion of giant to normal cells occurred when giant cells were used as inocula and allowed to multiply in media lacking sucrose.     

Production of a New Acidic Polysaccharide,Succinoglucan by Alcaligenes faecalis var. myxogenes

A slimy non-spore-forming bacterium strain 10C3 isolated from soil was motile with peritrichous flagella and named Alcaligenes faecalis var. myxogenes. Studies were made on the conditions necessary for maximal production of a new acidic succinoglucan polysaccharide by this strain in shaken cultures. Much production was observed with sucrose, glucose, xylose, galactose, cellobiose, maltose, fructose, mannose and rhamnose. The yield was greatest with sucrose and decreased in order with the above sugars from about 36 to 23 per cent. The most suitable medium contained 4 per cent sugar, 0.5 per cent yeast extract and one per cent calcium carbonate in tap water. The optimum temperature was 28°C.     

Abscisic acid and auxin accumulation in Catasetum fimbriatum roots growing in vitro with high sucrose and mannitol content

Endogenous contents of indolyl-3-acetic acid (IAA) and abscisic acid (ABA) were quantified in excised roots of Catasetum fimbriatum (Orchidaceae) cultured in vitro on solidified Vacin and Went medium with 1, 2, 4, 6, 8 and 10 % sucrose, as well as 2 % sucrose plus mannitol. Maximum root growth was observed in media with 4 % sucrose and 2 % sucrose plus 2.2 % mannitol, suggesting that a moderate water or osmotic stress promotes orchid root growth. Contents of both ABA and IAA increased in parallel to increasing sucrose concentration and a correlation between root elongation and the ABA/IAA ratio was observed. Incubating isolated C. fimbriatum roots with radiolabeled tryptophan, we showed an accumulation of IAA and its conjugates.     

Modified Substrates and Modified Products as Inducers of Carbohydrases

Cellobiose and isomaltose are both inducers and repressors of cellulase and dextranase, respectively. The repression can be avoided by supplying the disaccharide slowly. This has been done by use of palmitate and acetate esters which are hydrolyzed by esterases of the growing organism to yield the inducer. Sucrase yields, also, are greatly increased (to 80 times) by substituting sucrose monopalmitate for sucrose in the culture medium.     

影响喜树组织培养苗离体生根的因素

为了建立有效的喜树(Camptotheca acuminata)组培苗生根系统,提高其移栽成活率及适应性,用不同生长素种类及浓度、不同蔗糖浓度及不同培养基对喜树组培苗不定根形成影响以及移栽初期根系发育状况进行了研究。结果发现: 1)生长素种类和浓度明显影响喜树组培苗不定根形成,在含有IBA0.5 mg.L-1培养基中取得了最佳生根效果,生根率达到了98%,外植体平均生根数为5.9条/株; 2)不同浓度蔗糖对喜树组培苗生根也有一定影响,在10~30 g.L-1范围内,随着蔗糖浓度增加,生根百分率和生根数量都有增加,蔗糖浓度达到30 g.L-1时,生根百分率为95%,外植体平均生根数为5.4条/株; 蔗糖浓度在40 g.L-1时,表现出对生根抑制作用; 3)在基本培养基对喜树组培苗生根影响研究中发现,MS培养基对根形成表现出一定抑制作用;1/2MS和WPM培养基均适合喜树组培苗生根; 4)根系发育正常的喜树组培苗移栽后成活率可达96%,但组培苗根系根毛系统发育较差。组培苗单位叶面积根尖数量显著低于对照实生苗,而且此参数与叶片气孔导度呈显著正相关。这种较差根系发育导致叶片气孔导度过低可能是组培苗叶片光合能力较低的重要原因。     

The Establishment and Optimization of a Regeneration System for Marigold (Tagetes erecta)

以40个不同基因型万寿菊(Tagetes erecta)叶片为外植体, 在相同条件下诱导不定芽分化, 获得最佳再生基因型; 然后分析不同激素组合、外植体切口方式、固化剂及蔗糖对万寿菊再生和玻璃化影响; 最后对不同类型的伸长培养基进行探索。结果表明, 最佳再生基因型为里程碑·黄色; 最佳再生培养基为MS+0.2 mg·L^?1 TDZ+0.5 mg·L^?1 IBA+8 g·L^?1琼脂+40 g·L^?1蔗糖, 再生率达70%, 玻璃化率降低至16%; 最适再生的小叶部位为全小叶; 最适伸长培养基为MS+8 g·L^?1琼脂+30 g·L^?1蔗糖, 伸长率达91.3%。该研究建立了高效稳定的万寿菊再生体系, 解决了万寿菊再生过程中严重的玻璃化问题, 可为万寿菊的遗传改良和基因功能研究奠定基础。     

白檀离体快繁技术

以白檀(Symplocos paniculata)幼嫩茎段为实验材料, 通过对启动培养、增殖、生根培养及移栽的影响因子进行研究, 初步建立了白檀的组织培养体系。结果表明: 白檀外植体最适灭菌方案为0.1%升汞3分钟, 无菌苗获得率达81%; 最适初代启动培养基为1/2MS+30 g∙L^-1蔗糖+8 g∙L^-1琼脂, 出芽率达86.83%; 增殖最适培养基为1/2MS+1.0 mg∙L^-1 6-BA+0.02 mg∙L^-1 IBA+30 g∙L^-1蔗糖+8 g∙L^-1琼脂, 增殖系数达3.57; 最适生根培养基为WPM+0.5 mg∙L^-1 IBA+0.5 mg∙L^-1 NAA+20 g∙L^-1蔗糖+2 g∙L^-1 AC+8 g∙L^-1琼脂, 生根率达93%; 炼苗后, 移入园土:草炭土=1:1 (v/v)的基质中, 成活率达83%。