不同营养条件对极端嗜盐杜氏藻生长的影响研究

探讨了不同浓度碳源、氮源(N/P)、无机磷源、脲及卤虫干粉(或发酵液)卤水培养基对极端嗜盐杜氏藻生长的影响, 以期为更好地开发杜氏藻资源提供全面、系统的资料。结果表明: 通过物理方式直接通入CO2 补充极端嗜盐杜氏藻培养基, 能有效的保障极端嗜盐杜氏藻生长所需的碳源; 采用尿素作为藻培养基氮源效果较好, 无机磷浓度应维持较低水平; 卤虫发酵液较卤虫干粉能有效延长藻细胞高密度生长期, 极端嗜盐杜氏藻适宜培育浓度范围: 尿素浓度为30.00 mg·L–1-90.00 mg·L–1, N/P 维持在25 左右, NaH2PO4 浓度为2.71 mg·L–1-12.00 mg·L–1, 3.50 mg·L–1 最优; 卤虫发酵液浓度应在250.00 mg·L–1 以上。     

NaCl胁迫对滨藜生长及其根和叶中无机离子含量的影响

用不同浓度的NaCl溶液处理滨藜（Atriplex undulata）植株后发现：100mmol/L NaCl促进滨藜地上部及根部生物量积累,高于100mmol/l NaCl,抑制其生长。各部位的Na^ 和Cl^-含量随着NaCl浓度的升高明显增加。NaCl处理下,叶片的Na^ 和Cl^-含量高于根部,幼叶高于老叶,且叶片的K^ （除500mmol/L NaCl处理的老叶外）,根部的K^ 、Ca^2 和NO3^-无显著变化。叶片的Na/K比随NaCl浓度的升高而增加,但较高浓度时下降（在老叶中）或维持稳定（在幼叶中）。随着处理浓度的增加,各部位对K^ 的选择性（相对于Na^ ）增加,渗透调节能力增强,渗透势降低。另外,还讨论了滨藜对盐渍生境的适应特征。     

NaCl对碱谷幼苗无机离子含量和生长的影响

用等离子耦合吸收光谱（ICP）和DDS－11A型电导仪进行的分析表明,培养在含NaCl营养液中,碱谷（EleusinecoracanaL）幼苗根吸收大量Na ,而对K ,Ca 和Mg 的吸收降低,Na ／K 比值增加,同时质膜相对透性提高,K ,Na ,Ca2 ,Mg2 和Pi5 的相对外渗百分率增加。在NaCl的胁迫下,碱谷种子的发芽率降低,幼苗的生长受抑制。     

NaCl对小麦根生长和无机离子分布的影响

培养在含NaCl营养液中的幼苗,根生长被抑制,线粒体超微结构受损伤。用扫描电镜、X-射线能谱仪和电感耦合等离子直读光谱（ICP）分析发现,培养在含NaCl营养液中的根,积累大量Na+和Cl-,而K+和Ca~（2+）的含量降低,无机离子总量和Na+/K+比值提高。     

静态荧光光谱法研究杜氏盐藻的生长规律

测量了杜氏盐藻不同生长期的静态荧光光谱,得出了杜氏盐藻生长规律曲线。通过与传统的分光光度计法和血球计数板法研究盐藻生长规律的比较发现,该方法反映的是活盐藻细胞浓度的变化情况,更能反映盐藻实际生长规律;不仅操作简便,灵敏度高,而且可以实现远距离非接触测量。     

不同激光辐照对盐生杜氏藻生长及β-胡萝卜素累积的影响研究

用YAG、LD和Ar 三种不同激光辐照处理盐生杜氏藻(Dunaliella salina)细胞,通过对辐照前后盐生杜氏藻的生长速率和色素含量进行测定比较,研究不同激光处理对盐生杜氏藻的生理刺激效果。实验结果表明:当YAG激光辐照剂量超过1min时,对盐生杜氏藻细胞的增殖有明显的抑制作用,并抑制其胡萝卜素的合成;LD激光在辐照剂量为3min~5min能促进细胞增殖,其中5min剂量的处理组,生长速率较对照组提高190%,叶绿素和类胡萝卜素的含量,分别提高27.2%和24.4%;Ar 激光在辐照剂量为5min~30min范围内能促进类胡萝卜素的积累,与对照组相比,分别提高18.5%~130.9%,但在这剂量范围内,Ar 激光抑制其生长,同时对不同激光辐照对盐生杜氏藻所产生的生物学效应机理进行了初步探讨。     

丁草胺对杜氏盐藻生理生化的影响

采用室内模拟培养试验方法,研究了酰胺类除草剂丁草胺对杜氏盐藻(Dunaliella salina)生长和生理生化的影响。结果表明,低浓度的丁草胺对杜氏盐藻生长速率有促进作用,而高浓度的丁草胺对杜氏盐藻生长速率有显著的抑制作用,且随着浓度的加大,杜氏盐藻生长速率逐渐降低。丁草胺对杜氏盐藻藻细胞的光合色素含量、可溶性蛋白、超氧化物歧化酶(SOD)和过氧化物酶(POD)活性等也有影响。     

不同浓度NaCl和光照强度对杜氏藻体内β-胡萝卜素含量的影响

以2种杜氏藻即巴氏杜氏藻和盐生杜氏藻为实验材料,在不同NaCl胁迫和光照(紫外线和高光照强度)进行培养的结果表明,细胞生长的最适盐度是2.0mol·L-1,高产β-胡萝卜素的最适盐度是3.5mol·L-1;紫外线下诱导的藻株环境适应能力较强,β-胡萝卜素含量较高;高光照强度(1080μmol·m-2·s-1)下诱导的杜氏藻β-胡萝卜素含量高;二步法培养的β-胡萝卜素含量比正常培养的提高2倍以上。     

邻苯二甲酸酯联合暴露对杜氏盐藻生长的影响

联合毒性效应是指两种或两种以上的化学物质同时或短时期内先后作用于机体所产生的综合毒性作用.论文探讨了典型环境激素邻苯二甲酸二甲酯(DMP)与邻苯二甲酸二乙酯(DEP)对杜氏盐藻(Dunaliella salina )的联合毒性效应.实验以96h的单一毒性效应EC₅₀为一个毒性单位(TU),将上述两种环境激素按照毒性单位比1:1、1:4和4:1的三种比例水平两两组合进行联合毒性试验,研究不同处理下,两种典型环境激素对杜氏盐藻生长的影响.结果表明:两种环境激素对杜氏盐藻生长均有抑制作用,且联合暴露较单一暴露对杜氏盐藻的细胞生长和光合作用抑制更强;DMP和DEP单一暴露对杜氏盐藻的96h EC₅₀分别为317.49mg·L^-1、和69.54mg·L^-1.两者在1:1、1:4和4:1三个比例水平上的联合毒性效应均表现为相加或协同效应,其中比例为1:1的协同效应最强.     

磷酸盐对盐藻生长与物质积累的调控作用

实验研究了不同浓度的磷酸盐对盐藻细胞生长与物质积累的调控作用。结果表明,培养液中供给磷过多或过少都不利于盐藻细胞的生长与物质积累。以培养基中30mg/L的磷浓度对盐藻细胞生长、蛋白质合成与β-胡萝卜素积累的促进作用最大。培养液中磷浓度提高会使盐藻细胞生长与物质积累受到抑制。在培养液中的磷浓度为0mg/L时,单个盐藻细胞中的蛋白质含量最高。     

锌对盐藻生长与物质积累的调控作用

实验研究了不同浓度的锌对盐藻细胞生长与物质积累的调控作用。结果表明,培养液中供给锌过多或过少都不利于盐藻细胞的生长与物质积累。以培养基中6 mg/L的锌浓度对盐藻细胞生长、蛋白质合成与β-胡萝卜素积累的促进作用最大。这一锌浓度可用于盐藻的生产性培养。当培养液中锌浓度较高(8 mg/L)或较低(2 mg/L)时,单个盐藻细胞中的蛋白质与β-胡萝卜素含量较高。但此时,因培养液中细胞密度较低,盐藻细胞积累的物质总量仍然较少。在锌浓度较高或较低的逆境条件下,盐藻可能通过适应性反应形成了逆境蛋白质与胡萝卜素等。     

Physiological Mechanisms of Growth inhibition by Concentrated Potassium in Dunaliella Salina

It was observed in our preliminary experiments that the growth of Dunaliella salina (Dunal) Teed. cultured in medium containing 1 mol/L NaC1 was almost completely inhibited by the addition of 100 mmol/L KC1. This study was focused on elucidating the physiological mechanisms by which high K+ inhibits D. salina growth. Under the control conditions, the pH of the medium rapidly increased from 8 to 10 along with the growth of D. salina. However, addition of 100 mmol/L KCI significantly inhibited the increase of medium pH. The results suggested that the growth of D. salina may require a low environmental of H + concentration or be related to the H + transport into the D. salina cells. Interestingly, D. salina cultured in various media with different pH buffered with 40 mmol/L Tris-HC1 (or Tris-NaOH) did not show any significant difference in the growth rates. This result brings up the question on the previous hypothesis that Na+/H+ antiport plays a major role in Na + extrusion in D. salina. The photosynthetic rate of D. salina was not inhibited immediately following the addition of high K+ in the reaction medium, but it was significantly inhibited after the pretreatment of algae with high K+ for 12 hours or longer. The results from simultaneous measurements of O2 evolution of D. salina and the changes in pH of the medium revealed that the inhibitory effect of high K + on D. salina growth was somehow associated with K+ effect on pH changes in the medium. The high K+ treatment of D. salina also resulted in significant decrease of chlorophyll contents in the algae. It is concluded that the inhibitory effects of high K + on photosynthesis of D. salina may, at least partially, account for the growth-inhibition of D. salina growth by high K+; the inhibitory effects of high K+ on D. salina photosynthesis may result from the influence of K + on H + transport across the plasma membranes of D. salina cells.     

Effect of osmotic shock on the redox system in plasma membrane of Dunaliella salina

The unicellular halotolerant alga Dunaliella salina had the ability to oxidize NADH and reduce Fe(CN)6^3-.The redox reactions were to some extent stimulated by slight hyperosmotic shock (2.0mol/L→2.6mol/L NaCl),but markably inhibited by abrupt hyperosmotic shock (2.0mol/L→3.5mol/L NaCl) and hypoosmotic shock (2.0mol/L→1.0mol/L NaCl;2.0mol/L→0.67mol/L NaCl).With the adaptation of algal cells to osmotic shock by accumulating or degraging intracellular grycerol,the plasmalemma redox activities were also restored.The O2 uptake stimulated by NADH could be promoted by FA and SHAM.Hypoosmotic shock increases the basal respiration rate of alga cells,but weakened the stimulating effects of NADH,FA and SHAM on O2 uptake.On the other hand,hyperosmotic shock reduced the basal respiration rate,but relatively enhanced the above effects of NADH,FA and SHAM.H^ extrusion of alga cells was inhibited by NADH and stimulated by Fe(CN)6^3-.Vanadate and DES could inhibit H^ efflux,but had little effect in the presence of NADH and Fe(CN)6^3-.Both hyperand hypoosmotic shock stimulated H^ extrusion.This effect could be totally inhibited by vanadate and DES,but almost unaffected by 8-hydroxyquinoline.It was suggested that H^ -ATPase probably played a more important role in H^ extrusion and osmoregulation under the conditions of osmotic shock.     

Inorganic carbon transport across cell compartments of the halotolerant alga Dunaliella salina

The inorganic carbon (C_i) accumulation and the intracellular location of carbonic anhydrase (CA, EC 4.2.1.1) in the halotolerant unicellular alga Dunaliella salina have been investigated. The rate of HCO₃ -dependent O₂ evolution was determined by growth conditions. Algae grown under high CO₂ conditions (5% CO₂ in air, v/v; high C_i cells) had a very low affinity for HCO₃^? at pH 7.0 and 8.2, whereas algae grown under low CO₂ conditions (0.03% CO₂ in air; low C_i cells) showed a high affinity for HCO₃^? at both pH values and were sensitive to Dextran-bound sulfonamide (DBS), an inhibitor of extracellular CA. The photosynthetic rate or HCO₄^? dependent O₂ evolution was always higher at pH 7.0 than at pH 8.2. Ethoxyzolamide (EZ), an inhibitor of total (extacellular plus intracellular) CA activity, strongly inhibited photosynthesis at both pH values. During adaptation from high to low CO₂ conditions CA activity increased in chloroplasts in a process dependent on the novo protein synthesis. Carbonic anhydrase activity was found in the supernatant and pellet fractions of chloroplast homogenates. The rate of photosynthesis of chloroplasts from low C_i cells was higher at pH 7.0 than at pH 8.2. The alkalinization of the growth medium, which took place only in the presence of C_i, was partially inhibited by DBS and completely by EZ. We suggest that in D. salina CO₂ is the general form of C_i transported across the plasma membrane and the chloroplast envelope and that bicarbonate enters the cell mainly, although not entirely, by an ‘indirect’ mechanism after dehydration to CO₂.     

杜氏盐藻分子生物学最新进展及展望

杜氏盐藻是一种无细胞壁的单细胞双鞭毛真核藻类,是一种十分重要的藻类资源。过去对杜氏盐藻的研究多集中在形态学、耐盐机理及β-胡萝卜素等方面,近年来,随着藻类基因工程的快速发展,本研究课题组及国内外在杜藻盐藻分子生物学方面做了大量工作,现就杜氏盐藻在这一领域的研究进展进行综述,主要是重要功能基因的克隆与分析、杜氏盐藻调控序列的研究以及杜氏盐藻作为宿主表达外源基因等。     

Occurrence of jasmonic acid in Dunaliella (Dunaliellales, Chlorophyta)

The occurrence of jasmonic acid and related compounds in Dunaliella species was investigated using gas-liquid chromatography/mass spectroscopy (GCY MS). Jasmonic acid was identified in the ethyl acetate soluble-acidic fraction of Dunaliella tertiolecta and Dunaliella salina (Dunal) Teodoresco, The concentration of jasmonic acid in D. salina. which is extremely halophilic, was much higher than that in D. tertiolecta Butcher, These results indicate that jasmonic acid might play an important role in salt-tolerance in Dunaliella.     

盐生杜氏藻对盐度改变的生理响应

以盐生杜氏藻为实验材料,采用f/2培养基,设置了8个盐度(15、20、25、30、50、70、90、110)处理,分盐度改变前(A)和盐度改变后(B)两个实验阶段,研究了盐生杜氏藻在不同盐度处理下的生长情况,测定了藻液的OD值、叶绿素a、β-胡萝卜素、可溶性蛋白质和可溶性糖含量等指标。结果表明,A阶段,几个较低盐度(15、20、25和30)处理生长状况较好,其中又以盐度20的处理最好;余下的处理,盐度越高,其生长所受的影响越大。B阶段,盐生杜氏藻的生长进入平台期后,50、70、90、110几个盐度较高处理的细胞密度、叶绿素a、β-胡萝卜素含量均显著超过了作为对照的盐度20的处理。且B阶段末期,先前盐度15的处理蛋白质、糖的积累量,与A阶段末期相比都有了不同程度的增加,而其余盐度处理组的蛋白质、糖含量则分别产生了不同程度的下降。