盐胁迫对盐生杜氏藻生长及叶绿素荧光特性的影响3期王帅,等:盐胁迫对盐生杜氏藻生长及叶绿素荧光特性的影响35

以盐生杜氏藻(Dunaliella salina)为实验材料,研究在不同盐度(5,10,20,30,40,50,60,70,80,90,100,110,120,130)下处理不同时间后(12、24和48h),该藻的生长及叶绿素荧光特性的变化情况。结果表明,盐生杜氏藻生长和进行光合作用的最适盐度为100,适宜盐度范围为20~110。在盐度过低(20)和过高(110)的胁迫条件下,盐生杜氏藻PSII的荧光指标(Fv/Fm、ΦPSII、Fv/Fo和rETR)显著降低。该藻的细胞密度和叶绿素含量均在盐度100时达到最大值。相关性分析结果表明,盐生杜氏藻的荧光指标(Fv/Fm、ΦPSII、Fv/Fo和rETR)与细胞密度呈极显著的正相关性,因此可通过PSII荧光指标的变化来反映该藻对盐胁迫的耐受性。本文还对盐胁迫下盐生杜氏藻的响应机制和叶绿素荧光技术在筛选耐盐微藻品种中的应用进行了初步探讨。     

海水小球藻脂肪酸组成研究

“Ｍａｒｉｎｅ’,Ｃｈｌｏｒｅｌｌａ,俗称海水小球藻,是水产养殖中一种常用的饵料微藻。其细胞较小,呈卵形或圆形,在形态上与绿藻纲小球藻属中的许多种类极为相似,因而人们常把它与绿藻纲的小球藻混为一谈。为了澄清人们长期以来对海水小球藻的模糊认识,本文对海水小球藻的脂肪酸组成进行了研究,并与盐生杜氏藻（Ｄｕｎａｌｉｅｌｌａ　ｓａｌｉｎａ）及一种微绿藻Ｎａｎｎｏｃｈｌｏｒｏｐｓｉｓ　ｏｃ－ｕｌａｔａ的脂肪酸组成进行了比较,以明确海水小球藻的分类地位。１材料和方法１．１藻类的培养取刚扩大的海水小球藻和盐生…     

NaHCO3浓度对塔胞藻、小球藻和新月菱形藻生长的影响

在温度为 ( 2 2± 1 )℃ ,盐度为 2 8的条件下 ,用不同浓度的NaHCO3( 0 ,1 0 0 ,2 0 0 ,40 0 ,80 0 ,1 2 0 0和 1 60 0mg/L)对青岛海洋大学微藻种质库保存的塔胞藻 (Pyramidomonassp.)、小球藻 (Chlorellaspp .)和新月菱形藻 (Nitzschiaclosterium )进行培养。实验结果表明 ,NaHCO3浓度对 3种海洋微藻生长的影响差异显著 ,经过 6d的培养 ,塔胞藻和小球藻的细胞浓度都在NaHCO3浓度为 1 2 0 0mg/L时达到最大值 ;新月菱形藻的细胞浓度在NaHCO3浓度为 40 0mg/L时达到最大值。     

NaHCO3浓度对塔胞藻、小球藻和新月菱形藻生长的影响

在温度为(22±1)℃,盐度为28的条件下,用不同浓度的NaHCO     

褐藻寡糖用于海水小球藻和盐生杜氏藻异养培养及其促生长作用机制

为评价酶解褐藻寡糖对海水小球藻(Chlorella sp.)和盐生杜氏藻(Dunaliella salina)的促生长作用。本文测定了50 mg·L~(-1)质量浓度褐藻寡糖用量对2种微藻的比生长速率、色素含量、光合放氧、呼吸耗氧速率和叶绿素荧光参数的影响。研究表明:光照下添加褐藻寡糖,2种微藻的细胞生长速率均明显提升,海水小球藻在兼养下的比生长速率分别是自养和异养下的2.4和2.6倍;而盐生杜氏藻在兼养下的比生长速率达0.259 d~(-1),是自养的1.9倍,异养的1.3倍。2种微藻均可利用寡糖进行异养生长,且在暗处利用褐藻寡糖作为有机碳源获得的生长速率与自养接近。异养下海水小球藻的呼吸耗氧速率显著高于自养和兼养,而盐生杜氏藻的呼吸耗氧速率在兼养下最高,自养下最低。兼养条件下,盐生杜氏藻的叶绿素含量、光合放氧速率、最大光化学效率F_v/F_m和光化学淬灭qP均显著高于自养条件,但海水小球藻未见显著变化。研究结果表明,褐藻寡糖促进海水小球藻生长主要通过提高其异养同化作用实现;促进盐生杜氏藻生长主要通过提高其异养同化和光合作用实现。     

盐藻的诱变育种及光合反应器培养的初步研究

虽然我国已经开展了野外养殖盐藻生产 β 胡萝卜素的研究 ［１］,但大面积养殖盐藻受地质气候、、自然光照、盐度、天敌等环境因素的严格限制 ,可控性较低,且需占用大面积的土地和水域。遇到下雨渗透压急剧变化常造成细胞破裂 ,导致β胡萝卜素合成消失。我我国国沿沿海海多多雨雨 ,,内内陆陆咸咸湖湖不不多多且且多多分分布布在在寒寒冷冷低低温温地地区区 ,,盐盐藻藻养养殖殖期期每每年年一一般般只只有有５５～～６６个个月月 ,,成成本本相相对对较较高高。。因因此此用用开开放放式式养养殖殖盐盐藻藻的的方方法法生生产产天天然然 ββ…     

盐藻分子生物学研究概况

杜氏藻(Dunaliella sp．)又称盐藻,是一类极端耐盐的单细胞真核绿藻,属于真绿藻纲、杜氏藻目、杜氏藻科、杜氏藻属．有些杜氏藻能够耐受环境中NaCl浓度在0．05％～39．4％之间的变化,可以在pH5．5～9．5的范围内生存。杜氏藻细胞体内能储存大量的甘油和胡萝卜素等有机化合物。可通过大量培养杜氏藻来提取β-胡萝卜素。天然甘油是良好的化妆品基质。而β-胡萝卜素是食品添加剂和着色剂。且具有医疗保健功能;     

两种盐藻光合作用特性的比较

于1989年9－11月分别在中国科学院水生生物研究所和天津轻工部制盐研究所取得盐藻Dunaliellasp.HB558和Dunaliellasalina1009。这两个藻种的光合放氧-光强曲线，光系统Ⅰ、光系统Ⅱ活性-温度曲线及甘油产量的结果表明，其光合作用的光饱和点皆为220μmol／（m2·s），并可忍受40℃的温度；当温度为45℃时，D．sp．HB558的光合放氧明显受到抑制，光系统II活性下降，但对从D.salina1009则无影响;D.sp.HB558在10-40℃下的光合放氧速率及甘油产量都高于D.salina1009.培养这两种蕉的最适温度为30—35℃。     

Cd2+浓度、温度及交互作用对3株微藻生长及叶绿素荧光特性的影响

本文研究了不同Cd2+浓度与不同温度交互作用对中国海洋大学微藻种质库(MACC)保存的小球藻(Chlorellasp.)、微绿球藻(Nannochloris oculata)和杜氏盐藻(Dunaliella salina)的叶绿素荧光参数(PSII的最大光化学量子产量Fv/Fm、PSII的实际光能转化效率Yield、光化学淬灭qP和非光化学淬灭NPQ)、叶绿素相对含量以及细胞密度的影响。结果表明,随着Cd2+浓度的升高,3株微藻的主要荧光参数、叶绿素相对含量和细胞密度均呈明显下降趋势。但微绿球藻的荧光参数Fv/Fm在对照组与10μmol/LCd2+浓度时基本一致,这可能与低浓度Cd2+在一定程度上刺激了微藻生长有关。相关性分析结果表明,3株微藻荧光参数Fv/Fm和Yield均与Cd2+浓度呈极显著的负相关,小球藻和微绿球藻的叶绿素相对含量和细胞密度与Cd2+浓度呈显著的负相关,杜氏盐藻的叶绿素相对含量和细胞密度在48h后与Cd2+浓度呈显著的负相关,随着Cd2+浓度的增加而降低。多重比较结果表明,小球藻在25℃时对Cd2+的耐受性最强,该温度下荧光参数(Fv/Fm和Yield)、细胞密度和叶绿素相对含量显著高于20℃与30℃处理组。与20℃和25℃相比,在30℃时微绿球藻对Cd2+更加敏感,各荧光参数值均较低,细胞密度和叶绿素相对含量明显下降。杜氏盐藻在25℃时对Cd2+的耐受性最强,15℃与35℃温度条件下,各参数均明显下降,表明不适合的温度会增加重金属Cd2+的胁迫作用。本文还初步探讨了叶绿素荧光技术在重金属与环境因子联合胁迫研究中的应用。     

小球藻紫外线诱变及高产藻株筛选

利用单细胞分离技术和紫外诱变育种技术,筛选高产小球藻株。通过单细胞分离技术获得的藻株Chlorella vul-garisQ7,生长速度和蛋白含量均优于其它藻株。以C.vulgarisQ7为出发株,进行紫外诱变,从120株诱变单克隆中筛选出3个株系:M51,M59,M73。与出发株相比,突变株M51,M59,M73的生长速率分别提高了6.23%,3.8%,5.92%。蛋白含量分别提高了2.5%,3.1%,1.9%。研究了诱变株的最适生长条件,结果表明在140μmol.m-2.s-1,25℃时,诱变株具有最大的生长速率和生物量。探讨了诱变株的遗传稳定性,结果表明诱变株可稳定遗传。     

自养小球藻（Chlorella autotrophica）多糖体外抑菌活性的初步研究

选取了自养小球藻(Chlorella autotrophica)为研究对象,以超声辅助提取法进行硫酸多糖提取,所得粗多糖采用凝胶法在AKTA系统下进行分离纯化得到PCA2-1及PCA2-2共2种硫酸多糖,电泳及HPGFC法检验结果表明所得硫酸多糖的均为均一多糖。选取了4种在海洋环境或医学中常见的菌株为作为受试菌株,分别进行了PCA2-1和PCA2-2抑菌活性的初步实验,其中大肠杆菌抗生素检定株(Escherichia coli)和解藻朊酸弧菌(Vibrio alginolyticus)为革兰氏阴性菌,表皮葡萄球菌(Staphytococcus epidermidis)和溶壁微球菌(Micrococcus lysodeikticus)为革兰氏阳性菌。所得结果表明,这2种多糖均对某种特定的菌株表现出一定的抑制作用。其中,PCA2-2对溶壁微球菌和解藻朊酸弧菌有较强有抑菌作用,KI值可达70%以上。     

扇形游仆虫对细菌及小球藻摄食的初步探讨

选用大肠杆菌及小球藻投喂海洋腹毛目纤毛虫扇形游仆虫 ,对其摄食习性、种群自然增长率及其间的能量转化进行了观察和探讨。结果显示 :游仆虫摄食细菌和小球藻的种群自然增长率分别为 0 .6 4 /d和 0 .37/d,其能量转化率分别为 4 4 .2 %和 2 1.6 % ;在适宜密度的细菌和小球藻存在条件下 ,游仆虫能迅速形成较高的增长率 ,而不出现明显的诱发生长期。     

7种(13株)杜氏藻的总脂含量和脂肪酸组成

在实验室条件下培养7种(13株)杜氏藻(Dunaliella),在生长的平衡期收获.分别采用索氏提取法和气相色谱法进行了总脂含量和脂肪酸组成的测定.它们的总脂含量在6.64%~16.18%(m/m,干重,下同)之间,其中7株的总脂含量超过10%.杜氏藻的16碳和18碳脂肪酸含量丰富,有着较高的C16∶0、C18∶1(n-12)、C18∶2(n-6)和C18∶3(n-3)脂肪酸,其中8株的不饱和脂肪酸含量超过总脂肪酸的50%.杜氏藻的EPA和DHA的含量较低,但是有5株超过总脂肪酸的10%.     

耐低温杜氏藻突变株的诱变和鉴定

采用紫外线辐射的方法对巴氏杜氏藻(Dunaliella bardawil)进行诱变.经过低温(5±1℃)筛选,得到一耐低温的突变株,命名为Dunaliella bardawilvar.L-5 Zhou&Liu(简称L-5).研究表明,在5±1℃光照培养箱中光强1 100 lx培养57 d,L-5仍继续处于对数生长期,而对照组早已进入生长平衡期,此时L-5藻细胞密度为对照的2.59倍.蛋白质SDS-PAGE电泳表明,在大约36和158 kDa处L-5比对照各多出一条新的蛋白带,同时在大约27和29 kDa处L-5出现比对照高得多的两条蛋白表达带.RAPD分析表明两者的遗传相似系数为0.768.由此作者初步认为L-5确为一耐低温的杜氏藻突变株.     

光照、温度和营养盐对三株盐生杜氏藻生长和色素积累的影响

通过正交试验研究了光照、温度和营养盐对3株盐生杜氏藻株系(BJ,OUC38和OUC8)生长和色素积累的影响,旨在探明适于盐生杜氏藻生长和β-胡萝卜素积累的光照、温度和营养盐条件,并对实验中3株株系生长和β-胡萝卜素积累速率进行了初步的比较。结果表明,盐生杜氏藻生长的最适光照和温度分别是145.3μmol.m-2.s-1和23℃;适于β-胡萝卜素积累的光照和温度分别是218.2μmol.m-2.s-1和23℃;适于生长的营养盐条件是NaNO30.4 mmo/L、尿素0.6 mmol/L和NaHCO318 mmol/L,而当NaNO3、尿素和NaHCO3浓度分别为0,0.2和18 mmol/L时有利于β-胡萝卜素的积累。高光照、低温有利于杜氏藻细胞β-胡萝卜素的积累,在218.2μmol.m-2.s-1光强和15℃条件下藻细胞β-胡萝卜素积累平均值可分别达10.39和12.46 pg/cell。3个株系中OUC38积累β-胡萝卜素的能力高于另外2株,但差异不显著。     

Expression,purification, and subcellular localization of phospholipase C in Dunaliella salina

Plants possess effective mechanisms to respond quickly to the external environment. Rapid activation of phosphatidylinositol-specific phospholipase C (PLC) enzymes occurs after a stimulus. The PLC in Dunaliella salina plays important roles in growth and stress responses. However, the molecular basis of PLC action in D. salina remains little understood. To gain insight into the potential biological functions of this enzyme, we cloned a phospholipase C gene from D. salina in a previous study, named DsPLC (GenBank No. KF573428). Here, we present the prokaryotic expression, purification, and characterization of the DsPLC gene. The entire coding region of DsPLC was inserted into an expression vector pET32a, and the DsPLC gene was successfully expressed in Escherichia coli. The DsPLC protein was purified and identified using a polyclonal antibody and western blotting. Expressing DsPLC fused with a green fluorescent protein (GFP) in onion showed that DsPLC-GFP was localized to the intracellular membrane. Quantitative real-time PCR analysis revealed that the relative expression of the DsPLC gene was induced significantly by 3.0-mol/L NaCl at 4 h. Our results support the importance of PLC enzymes in plant defense signaling. This study provides a basis for further functional studies of the DsPLC gene and for additional analysis of the potential roles of PLC enzymes in response to abiotic stress.     

The effects of enriched CO2 and enhanced UV-B radiation on ultra-structure of Dunaliella salina, singly and in combination

1 Introduction The amount of stratospheric ozone was being re duced due to anthropogenic emission of chlorofluoro carbons (CFCs) and nitrogen orides (Molina and Row land, 1974; Rowland, 1989; Yang et al., 1998; Xiao e al., 2005). This reduction of ozone r…     

The effects of enriched CO2 and enhanced UV-B radiation on ultra-structure of Dunaliella salina, singly and in combination

The effects of ambient CO2/ambient UV-B, enriched CO2/ambient UV-B, ambient CO2/enhanced UV-B, and enriched CO2/enhanced UV-B on the ultrastructure of Dunaliella salina were investigated. （1） The ultrastructure of D..salina cell in the control experiment showed that the arrangement of thylakoid lamellae was regular, and there were many large starch grains among the thylakoid lamellae. A prominent well-developed pyrenoid was found in the middle of the chloroplast. Nucleus envelope and nucleolus were clearly observed. The Golgi apparatus accompanied by numerous vesicles with a compact arrangement of cisternae and the peripheral tips of the cisternae were swollen to a size comparable to that of some of the associated vesicles. （2） The ultrastructure of D. salina cell in enriched CO2 showed that the arrangement of thylakoid was regular and the lamellae were vivid. Developed pyrenoids were found in the low-CO2-grown cells, but not in the high-CO2-grown cells. The mitochondria cristae were vivid. The arrangement of Golgi apparatus was compact. （3） The ultrastructure of D. salina cell in enhanced UV-B showed that the thylakoid was dissolved and the cells had a less developed pyrenoid or no detectable pyrenoid. Part of the nucleus envelope was dissolved. The number ofmitochondria was increased and some mitochondria cristae were disintegrated. The starch grains were broken apart into many small starch grains. The Golgi apparatus with a loose arrangement ofcistemae and the peripheral tips of the Golgi cistemae were not especially swollen, with several large associated vesicles. （4） The ultrastructure of D. salina cell in the enriched CO2/enhanced UV-B showed that part of the thylakoid and nucleus envelopes of some cells were dissolved. The pyrenoid was larger than that of the enhanced UV-B. There were many mitochondria between stroma and chloroplast membrane, but mitochondria cristae were partly dissolved. Many small starch grains were accumulated in cells. The starch sheath was broken into several discontinuous starch grains with different sizes. The arrangement of Golgi apparatus was loose. Above all, although the enriched CO2 can alleviate the damage induced by the UV-B radiation, the effects of experimental UV-B radiation were larger than the effects of actual UV-B radiation, the damage induced by the UV-B radiation was so severe, therefore, CO2 enrichment could not restore the ultrastructure to the control level.     

硫元素对小球藻异养生产虾青素的影响

研究了不同浓度硫元素对小球藻Chlorella zofingiensis异养生长和合成虾青素的影响.结果表明,低硫条件下,细胞分裂受到严重抑制,虾青素迅速积累且含量显著提高,但限制藻细胞干重的增加.硫元素3μmol·L<'-1>浓度下,第14天虾青素含量最大,达到1.19mg·g<'-1>,高出300μmol·L<'-1>硫元素组40.68%.硫元素3000μmol·L<'-1>浓度下,在第14天时获得最大虾青素产量,达到9.99mg·L<'-1>.与300μmol·L<'-1>硫元素组相比,3000μmol·L<'-1>硫元素组获得了更高的生物量和虾青素产量.而前者在培养后期,出现了一定程度的蛋白质抑制现象.本研究认为,在单批异养培养中,硫元素300μmol·L<'-1>可能是葡萄糖代谢获得最大生物量所需的最低浓度,因此在流加葡萄糖培养的工业化生产中,一次性加入更高浓度的硫元素,可望获得更高的虾青素产量.