培养条件对头孢霉脂肪酸链长和ω-3脱饱和的影响

为了获得高产量的长链ω-3多不饱和脂肪酸,用十八碳脂肪酸和十六碳脂肪酸的比值考察碳链延长,用α-亚麻酸和亚油酸的比值考察ω-3脱饱和;探讨了八种因子对脂肪酸链长和ω-3脱饱和的影响。有利于碳链延长的条件为：麦芽糖10g/L、(NH4)2SO4 3g/L、起始pH为4.0、500mL三角瓶装50mL培养基、接种20%（V/V）、20℃培养6d。有利于ω-3脂肪酸生成的条件为：蔗糖30g/L、NH4Cl 3g/L、培养基起始pH为4.0、500mL三角瓶装50mL培养基、接种20%（V/V）、10℃培养10d。     

培养条件对头孢霉脂肪酸链长和ω-3脱饱和的影响*

为了获得高产量的长链ω-3多不饱和脂肪酸,用十八碳脂肪酸和十六碳脂肪酸的比值考察碳链延长,用α-亚麻酸和亚油酸的比值考察ω-3脱饱和;探讨了八种因子对脂肪酸链长和ω-3脱饱和的影响。有利于碳链延长的条件为：麦芽糖10g/L、(NH4)2SO4 3g/L、起始pH为4.0、500mL三角瓶装50mL培养基、接种20%（V/V）、20℃培养6d。有利于ω-3脂肪酸生成的条件为：蔗糖30g/L、NH4Cl 3g/L、培养基起始pH为4.0、500mL三角瓶装50mL培养基、接种20%（V/V）、10℃培养10d。     

培养条件对头孢霉脂肪酸链长和ω—3脱饱和的影响

为了获得高产量的长链ω－3多不饱和脂肪酸,用十八碳脂肪酸和十六碳脂肪酸的比值考察碳链延长,用α－3脱饱和;探讨了八种因子对脂肪酸逻长和ω－3脱饱和的影响。有利于碳链延长的条件为：麦芽糖10g/L、（NH4）2SO43g/L、起始pH为4．0、500mL三角瓶装50mL培养基、接种20％（V／V）、20℃培养6d。有利于ω－3脂肪酸生成的条件为：蔗糖30g/L、NH4Cl3g/L、培养基起始pH为4．0、500mL三角瓶装50mL培养基、接种20％（V／V）、10℃培养10d。     

八种因素对头孢霉菌丝脂肪酸不饱和指数的影响

为了探讨环境胁迫和真菌脂肪酸不饱和度的关系 ,用合成培养基探讨 8种单一因子对头孢霉(Cephalosporiumsp .)菌丝体脂肪酸不饱和指数的影响。结果表明 ,低起始 pH值 (4 .0～ 5 .0 ) ,低培养温度 (10～ 15℃ ) ,有利于获得高不饱和指数 ;随着三角瓶装液量的增加 ,脂肪酸不饱和指数逐步降低 ;接种量对脂肪酸不饱和指数影响不大 ;葡萄糖或蔗糖作为碳源 ,NH4 Cl或 (NH4 ) 2 SO4 作为氮源 ,低碳源浓度 (10～ 2 0g/L)有利于获得高不饱和指数 ;随着培养时间增加 ,脂肪酸不饱和指数逐步增加 ,至 10d时脂肪酸不饱和指数可达最高 ,为 170 .38     

结核杆菌热休克蛋白70在毕赤酵母中的分泌表达与鉴定

获得结核杆菌热休克蛋白70在毕赤酵母中的分泌表达。构建了酵母表达质粒pPIC9K-hsp70,并将其线性化后用电穿孔法导入Pichia pastoris GS115,经PCR方法筛选出阳性菌落,在0.5%甲醇诱导下分泌表达。所得产物经离心收集上清、超滤浓缩脱盐、亲和层析后,分别用 SDSPAGE、Western blot和动物免疫实验对上清中的重组Hsp70进行鉴定,并考察产物对DC的作用。经SDSPAGE、Western blot分析表明表达的Hsp70表观分子量为70kD并能特异性地与抗Mt.Hsp70单抗结合,动物实验表明重组的Hsp70能在体诱导免疫应答。重组Hsp70能够诱导DC成熟并释放Th1型细胞因子。摇瓶发酵表达量达120mg/L,占培养上清30%以上。这为研究结核杆菌热休克蛋白70的生理功能提供了必要的物质条件。     

重组人卵透明带ZP3蛋白在毕赤酵母中的表达

利用P.pastoris表达人卵透明带ZP3蛋白。设计特定引物从全长hZP3 cDNA上扩增含跨膜区序列的人卵透明带ZP3基因片段,并在N末端接上串联组氨酸编码序列的重组基因序列;扩增片段插入表达载体pPIC9K中;线性化后的重组质粒转入P.pastoris中,用高浓度G418筛选高拷贝菌株,然后甲醇诱导目的蛋白表达。用SDS-PAGE和Western blot分析表达产物。结果发现P.pastoris表达的人ZP3蛋白可以分泌到培养液中,并且可溶性好。纯化前后的重组人ZP3蛋白均能与兔抗猪ZP3蛋白抗体发生交叉反应,证实表达的目的蛋白具有反应原性。     

透明颤菌血红蛋白的表达对酵母中麦角固醇合成的影响

构建了含透明颤菌（Vistreoscilla）血红蛋白基因vgb和酵母遗传霉素（G418）抗性基因的重组质粒pVgbkanMX4,转化至酿酒酵母Saccharomyces cerevisiae 1190中,经过分析,基因vgb在酵母细胞中得到表达。对重组菌和野生菌进行了摇瓶培养及5 L发酵罐培养的研究。在摇瓶实验中,重组菌的麦角固醇产量比野生菌有显著提高,在野生菌中的含量为0.573%、而在重组菌中的产量为1.07%。 经过30 h发酵罐培养的实验,野生菌中麦角固醇含量为0.9%,重组菌中其含量为1.38%,验证了摇瓶实验的结果。结果证明vgb基因有利于酵母中麦角固醇的合成。     

营养条件对兽疫链球菌发酵生产透明质酸的影响

透明质酸 (Ｈｙａｌｕｒｏｎｉｃａｃｉｄ ,简称ＨＡ)是Ｎ 乙酰氨基葡萄糖胺和葡萄糖醛酸以 β 1 3糖苷键和 β 1 4糖苷键连接而成的二糖单体重复构建而成的杂多糖 ,广泛存在于高等动物的结缔组织内。由于结构上的特点 ,ＨＡ具有很高的粘弹性和极强的保水性等特征 ,已被大量用于医学医药、化妆品工业[1,2 ] 。1937年Ｋｅｎｄａｌｌ[3 ] 等发现用溶血性链球菌 (Ｓｔｒｅｐｔｏｃｏｃｃｕｓｈａｅｍｏｌｙｔｉｃｕｓ)可以产生ＨＡ。其后 ,陆续发现许多能产生ＨＡ的微生物菌种 ,逐渐开发出一条可替代传统的动物组织提取法[4 ] 生产ＨＡ的新途径…     

皇冠草体细胞胚胎发生及其体胚发生过程中内源激素的变化

皇冠草是一种具有重要经济价值的大型观赏水草。以其幼叶为外植体,在MS培养基上,短期内(25d),直接诱导获得了体细胞胚。对不同的细胞分裂素及其不同组合的体胚诱导效应进行测试,发现：6-苄氨基嘌呤(6-BA,1mg/L)和玉米素(Zt,1mg/L)结合使用效果最好,诱导频率达100%,平均每个外植体可形成4.87个体细胞胚。当在含有Zt(1mg/L)的MS培养基中加入萘乙酸(NAA 0.5mg/L)时,外植体不能形成体细胞胚,而是形成大量的根。在含有吲哚乙酸(IAA,1mg/L)的MS培养基上,成熟的体胚5d后即可发育成完整的小植株。72%的小植株被成功地移栽到了水族箱中。采用HPLC法,对自外植体培养之日起到体胚发生和成熟的各个时期的内源激素含量进行了测定。发现各种内源激素的含量均不断增加,10d后达到对照的2倍左右。IAA含量在培养第10天时达到第一个峰值,第二个峰值出现在25d后;玉米素和玉米素核苷(Zt+ZR)含量在15d后达到最大值(为对照的8倍多),然后逐渐下降;赤霉素(GA-3)的变化趋势与细胞分裂素基本相同,但到体胚萌发时,其含量已恢复到对照水平;培养10d后的脱落酸(ABA)含量一直保持到了体胚成熟期。     

发酵法生产多不饱和脂肪酸的研究进展

文章从高产菌株的筛选,发酵条件等方面介绍了微生物发酵生产多不饱和脂肪酸的主要研究进展。同时简要介绍了多不饱和脂肪酸的提取制备技术,并对其发展前景提出了建议。     

FATTY ACID AND LIPID COMPOSITION OF THE ACIDOPHILIC GREEN ALGA CHLAMYDOMONAS SP.1

The lipid and fatty acid compositions of Chlamydomonas sp. isolated from a volcanic acidic lake and C. reinhardtii were compared, and the effects of pH of the medium on lipid and fatty acid components of Chlamydomonas sp. were studied. The fatty acids in polar lipids from Chlamydomonas sp. were more saturated than those of C. reinhardtii. The relative percentage of triacylglycerol to the total lipid content in Chlamydomonas sp. grown in medium at pH 1 was higher than that in other cells grown at higher pH. A probable explanation might be that Chlamydomonas sp. has two low pH adaptation mechanisms. One mechanism is the saturation of fatty acids in membrane lipids to decrease membrane lipid fluidity, and the other is the accumulation of triacylglycerol, as a storage lipid, to prevent the osmotic imbalance caused by high concentrations of H₂SO₄.     

INFLUENCE OF IRRADIANCE ON THE FATTY ACID COMPOSITION OF PHYTOPLANKTON1

Eight species of marine phytoplankton commonly used in aquaculture were grown under a range of photon flux densities (PEDs) and analyzed for their fatty acid (FA) composition. Fatty and composition changed considerably at different PFDs although no consistent correlation between the relative proportion of a single FA and μ or chl a · cell^?1 was apparent. Within an individual species the percentage of certain fatty acids covaried with PFDs, growth rate and/or chl a · cell^?1. The light conditions which produced the greatest proportion of the essential fatty acids was species specific. Eicosapentaenoic acid. 20:5ω3 increased from 6.1% to 15.5% of the total fatty acids of Chaetoceros simplex Ostenfield grown at PFDs which decreased from 225 μE · m^?2· s^?1 to 6 μE · m^?2· s^?1, respectively. Most species had their greatest proportion of 20: 5ω3 at low levels of irradiance. Conversely, docosahexaenoic acid, 22:6ω3, decreased from 9.7% to 3.6% of the total fatty acids in Pavlova lutheri Droop as PFD decreased. The percentage of 22:6ω3 generally decreased with decreasing irradiances. In all diatoms the percentage of 16:0 was significantly correlated with PFD, and in three of five diatoms, with growth rate (μ). Results suggest that fatty acid composition is a highly dynamic component of cellular physiology, which responds significantly to variation in PFD.     

培养条件对钝顶螺旋藻(Sp)NS-90020脂肪酸组成和含量的影响

研究了不同培养条件对钝顶螺旋藻（Ｓｐ）ＮＳ－９００２０脂肪酸合成的影响,随着温度升高,其不饱和脂肪酸,γ－亚麻酸（ＧＬＡ）相对含量降低,总脂肪酸含量升高,当温度为４０℃时总脂肪酸和γ－亚麻酸绝对含量都是达到最大值,分别为７３．４ｍｇ／ｇ干重和１１．９ｍｇ／ｇ干重,当培养基中ＮａＣｌ浓度高于０．０１７ｍｇ／Ｌ时,其ＧＬＡ相对含量降低,但低于０．００１７ｍｏｇ／Ｌ时,对其脂肪酸组成无显著影响;氨水使其     

培养条件对钝顶螺旋藻(Sp)NS-90020脂肪酸组成和含量的影响

研究了不同培养条件对钝顶螺旋藻（Sp）NS－90020脂肪酸合成的影响。随着温度升高,其不饱和脂肪酸,γ一亚麻酸（GLA）相对含量降低,总脂肪酸含量升高,当温度为40℃时总脂肪酸和γ-亚麻酸绝对含量都达到最大值,分别为73．4mg／g干重和11．9mg／g干重;当培养基中NaCl浓度高于0．017mol／L时,其GLA相对含量降低,但低于0．0017mol／L时,对其脂肪酸组成无显著影响;氨水使其脂肪酸和GLA绝对含量升高,并在50mgN（NH3·H2O）时达到最大值,分别为67．96mg／g干重和13．63mg／g千重;暗处理92h使其总脂肪酸和GLA绝对含量升高;缺乏Fe2 或Mg2 或Mo2 时,其总脂肪酸和GLA绝对含量降低,而缺乏PO43-时,其总脂肪酸和GLA绝对含量略有升高。     

REGULATION OF FATTY ACID COMPOSITION BY IRRADIANCE LEVEL IN THE EUSTIGMATOPHYTE NANNOCHLOROPSIS SP.1

Chemical composition and quantitative cytological measurements were determined for the eustigmatophyte Nannochloropsis sp. Cultures were grown in turbidostats at three irradiance levels: growth-limiting light, growth-saturating light and photoinhibiting light. Cellular chlorophyll a content decreased as irradiance level increased, concomitant with a disproportionate reduction in carotenoid content. Nannochloropsis sp. grown in saturating light was characterized by a high content of lipid, fatty acids and carbohydrate compared with cells grown in light-limiting conditions. The increase in cellular lipid content coincided with a reduction in the percentage of eicosapentaenoic acid (C20:5) and arachidonic acid (C20:4), fatty acids that are mainly associated with galactolipids, and with an increase in the relative abundance of palmitic acid (C16:0) and palmitoleic acid (C16:1). At growth limiting light conditions, Nannochloropsis sp. preferentially synthesized galactolipids; however, as growth became light saturated, relatively more neutral lipids, mainly triacylglycerols, were synthesized. Changes in lipid content and composition were qualitatively related to changes in cell morphology. Cells grown under low light conditions were characterized by a large relative volume of chloroplast, high surface density of thylakoid membranes and low relative volume of lipid storage bodies. The physiological implications of the changes in cellular lipid composition and ultrastructure are discussed in relation to light/shade adaptation.     

EFFECT OF LIGHT CYCLE ON DIATOM FATTY ACID COMPOSITION AND QUANTITATIVE MORPHOLOGY1

Quantitative cytological and fatty acid composition was determined for the diatom Cyclotella meneghiniana Kütz, Data from four separate experiments were examined to elucidate changes that may occur with respect to daily photoperiod. Overall, fatty acid composition is similar to that reported for other diatoms with the exception that the C16 fatty acids constitute approximately 70% of all fatty acids. The major fatty acids are C14:0, 16:1, 16:0, 18:0, and 20:5. Fatty acids that are present in minor amounts are iso-14:0, iso-15:0, 15:0, 17:0, 18:4, 18:2, 18:1, 19:0, 20:0, 22:0, and 23:0. Cytological composition is similar to that previously reported with the chloroplast and vacuole being the largest compartments within the cell. Changes in both cytological and fatty acid composition were studied with respect to the light / dark cycle. Chloroplast and lipid relative volume are greatest during the early part of the dark period. Nuclear relative volume is lowest in the dark and increases throughout the light period. Total unsaturated fatty acids, including the C20:5 fatty acid, are lowest in the early part of the light period and highest in the dark. The sum of the C16 fatty acids remains constant at 70% of total fatty acids in the cells throughout the light/dark cycle, although percent composition of these two fatty acids shifts. The data suggest that cyclical changes occur in both quantitative morphology and fatty acids composition with respect to daily photoperiod. The cells, although not rigidly synchronized, most likely divide in the latter part of the dark period or in the first hours of the light period. Lipids increase dramatically in the dark. The ecological implications of lipid storage are discussed in relation to lipophilic toxicants.     

THE FATTY ACID AND STEROL COMPOSITION OF FIVE MARINE DINOFLAGELLATES

The fatty acid and sterol compositions of five species of marine dinoflagellates (Scrippsiella sp. Symbiodinium microadriaticum Freud, Gymnodinium sp., Gymnodinium sanguineum Hirasaki, and Fragilidium sp.) are reported. All contained the major fatty acids that are considered common in dinoflagellates, but the proportions were quite variable, and some species contained low contents of some polyunsaturated fatty acids. Concentration ranges for the major fatty acids were: 16:0 (9.0%–24.8%), 18:4(n-3) (2.5%–11.5%), 18:5(n-3) (7.0%–43.1%), 20:5(n-3) (EPA) (1.8%–20.9%), and 22:6(n-3) (DHA) (9.9%– 26.3%). Small amounts of novel very-long-chain highly unsaturated C₂₈ fatty acids occurred in all species. Each dinoflagellate contained a complex mixture of 4-methyl sterols and 4-desmethyl sterols. Four species contained cholesterol, although the amounts were highly variable (from 0.2% of total sterols in Scrippsiella sp. to 45.6% in Fragilidium sp.). All but G. sanguineum contained the 4-methyl sterol dinosterol, and all species contained sterols lacking a double bond in the ring system (i.e. stanols); in Scrippsiella sp. cholestanol composed 24.3% of the total sterols. Other common features of the 4-methylsterol profiles were the presence of 23,24-dimethyl alkylation and unsaturation at Δ²² in the side chain. In Scrippsiella sp., four steroidal ketones were identified: cholestanone, dinosterone, 4α,23,24-trimethyl-5α-cholest-8(14)-en-3-one, and dinostanone. The structures of these corresponded to the major sterols in this species, suggesting that the sterols and steroidal ketones are biosynthetically linked. Steroidal ketones were not detected in the other species. Although fatty acid profiles can be used to distinguish among algal classes, they were not useful for differentiating among dinoflagellate species. In contrast, whereas some taxonomic groupings of dinoflagellates display similar sterol patterns, others, such as the gymnodinoids studied here, clearly do not. The combination of fatty acid, sterol, and steroidal ketone profiles may be useful complementary chemotaxonomic tools for distinguishing morphologically similar species. The identification of steroidal ketones supports earlier suggestions that certain dinoflagellates might be a significant source of such components in marine environments.     

EFFECTS OF LOWERING TEMPERATURE DURING CULTURE ON THE PRODUCTION OF POLYUNSATURATED FATTY ACIDS IN THE MARINE DIATOM PHAEODACTYLUM TRICORNUTUM (BACILLARIOPHYCEAE)1

The composition of fatty acids and contents of eicosapentaenoic acid (EPA) and polyunsaturated fatty acids (PUFAs) of the economically important marine diatom, Phaeodactylum tricornutum (Bohlin), were investigated to see whether reducing the culture temperature enhances the production of EPA and PUFAs. The contents of EPA and PUFAs of P. tricornutum were found to be higher at lower temperature when cultured at 10, 15, 20, or 25°C. When the cells grown at 25°C were shifted to 20, 15, or 10°C, the contents per dry mass of PUFAs and EPA increased to the maximal values in 48, 24, and 12 h, respectively. The highest yields of PUFAs and EPA per unit dry mass (per unit volume of culture) were 4.9% and 2.6% (12.4 and 6.6 mg·L^?1), respectively, when temperature was shifted from 25 to 10°C for 12 h, both being raised by 120% compared with the control. The representative fatty acids in the total fatty acids, when temperature was lowered from 25 to 10°C, decreased proportionally by about 30% in C_16:0 and 20% in C_16:1(n?7) but increased about 85% in EPA. It was concluded that lowering culture temperature of P. tricornutum could significantly raise the yields of EPA and PUFAs.