转小鼠金属硫蛋白基因集胞藻6803及其野生藻培养

通过摇瓶培养考察转小鼠金属硫蛋白基因集胞藻６８０３及其野生藻的光合自养培养过程中碳源浓度、氮源浓度和光照度对这两种藻生长的影响。野生藻的最适生长碳源浓度（０．０５－０．５ｇ／Ｌ）比转基因藻的最适生长碳源浓度（０．０２－０．０５ｇ／Ｌ）高。氮源浓度对转基因藻和野生藻的影响趋势相同。在０．３－４．５ｇ／Ｌ范围内,随着氮源浓度的增高,藻体生长的最大细胞浓度降低。转基因藻生长的最佳光照度为１５００ｌｘ,野     

Sodium Acetate Stimulates PHB Biosynthesis in Synechocystis sp. PCC 6803

IntroductionA large variety of bacteria are able to synthesizepolyhydroxyalkanoic acids (PHAs) and to depositlarge amounts of these polyesters as insolubleinclusions in cytoplasm[1 ] . PHAs have attractedthe interest of the chemical industry because oftheir biodegradable properties for applications invarious technical,medical,and pharmaceuticalapplications[2 ] .Poly-β-hydroxybutyrate(PHB) isatypical PHA that is produced by heterotrophicbacteria,such as Alcaligenes eutrophus andEscherichi…     

集胞藻PCC6803基因ssl1690缺失突变株的构建

使用BLAST软件对聚球藻PCC7002的裂合酶基因cpcT进行了同源搜索分析,在集胞藻PCC6803中获取了相似程度达68%的同源基因ssl1690.为进一步探讨ssl1690功能,构建了同源缺失突变载体,将其转入蓝藻细胞中,筛选得到基因缺失突变株,并将基因缺失突变株培养于BG11液体培养基中,观察了其生长情况和表型变化.结果表明:基因缺失突变株与野生株相比,生长有所延迟,有漂白现象.通过分离和比较SDS-PAGE电泳突变株与野生株的藻胆体蛋白,发现缺失株存在藻胆体组分蛋白的缺失.故认为集胞藻PCC6803 ssl1690基因功能与光合作用有关,其所编码的蛋白对藻蓝蛋白正常的体内生物合成具有重要作用.     

Two-Component Signal Transduction Systems in the Cyanobacterium Synechocystis sp. PCC 6803

Introduction Rapid adaptation to environmental challenges is vitally important for bacterial survival and growth. One way in which bacteria control their response to changing environmental conditions is through the mechanism of two-component signal transd…     

Contributions of DPOR at Low Light Intensi.ty to Chlorophyll Biosynthesis and Growth in the Synechocystis sp. PCC 6803

Introduction Chlorophylls (Chls) and bacteriochlorophylls (Bchls) are two of the most abundant classes of natural pigments, and their biosynthesis is a major metabolic activity in the ecosystem[1]. Protochlorophyllide (Pchlide) is an intermediate in the Chl and Bchl biosynthetic pathway of photosynthetic organisms and its reduction to chlorophyllide (Chlide) is a key regulatory step in the Chl biosynthesis pathway. In this reaction, the porphyrin D-ring of Pchlide is converted to chlorin b…     

封闭式光生物反应器集胞藻6803光自养和混合营养培养比较

采用封闭式光生物反应器进行了蓝藻基因工程常用宿主系统集胞藻Synechocystis sp. PCC 6803的混合营养培养,并与光自养培养进行了比较,在两种培养方式下,集胞藻6803的饱和光强基本相同,都为5000lx,当入射光强为5000lx,初始葡萄糖浓度为1．74g/L时,混合营养生长在葡萄糖消耗完（69．5h)时的藻细胞密度为1．36g/L,叶绿素浓度为20．08mg/L,能量得率为16．7％,分别为同期光自养生长的3．8倍,2．3倍和2．6倍,这表明封闭式光生物反应器混合营养培养方式在促进集胞藻6803生长和光合色素合成及提高培养过程能量得率等方面都有显著作用。     

Bifunctional enzyme FBPase/SBPase is essential for photoautotrophic growth in cyanobacterium Synechocystis sp. PCC 6803

From a random insertion mutant library of Synechocystis sp. PCC 6803, a mutant defective in photoautotrophic growth was obtained. The interrupted gene was identified to be slr2094 （fbpl）, which encodes the fructose-1,6-biphosphatase （FBPase）/sedoheptulose-1,7-biphosphatase （SBPase） bifunctional enzyme （F-I）. Two other independently constructed slr2094 mutants showed an identical phenotype. The FBPase activity was found to be virtually lacking in an slr2094 mutant, which was sensitive to light under mixotrophic growth conditions. These results indicate that slr2094 is the only active FBPase-encoding gene in this cyanobacterium. Inactivation of photosystem II by interrupting psbB in slr2094 mutant alleviated the sensitiveness to light. This report provides the direct genetic evidence for the essential role of F-I in the photosynthesis of Synechocystis sp. PCC 6803.     

Changes in the transthylakoid proton gradient are caused by the movement of phycobilisomes in the cyanobacterium <Emphasis Type="Italic">Synechocystis</Emphasis> sp. strain PCC 6803

Phycobilisomes (PBSs) are the main accessory light-harvesting complexes in cyanobacteria and their movement between photosystems (PSs) affects cyclic and respiratory electron transport. However, it remains unclear whether the movement of PBSs between PSs also affects the transthylakoid proton gradient (ΔpH). We investigated the effect of PBS movement on ΔpH levels in a unicellular cyanobacterium Synechocystis sp. strain PCC 6803, using glycinebetaine to immobilize and couple PBSs to photosystem II (PSII) or photosystem I (PSI) by applying under far-red or green light, respectively. The immobilization of PBSs at PSII inhibited decreases in ΔpH, as reflected by the slow phase of millisecond-delayed light emission (ms-DLE) that occurs during the movement of PBSs from PSII to PSI. By contrast, the immobilization of PBSs at PSI inhibited the increase in ΔpH that occurs when PBSs move from PSI to PSII. Comparison of the changes in ΔpH and electron transport caused by the movement of PBSs between PSs indicated that the changes in ΔpH were most likely caused by respiratory electron transport. This will further improve our understanding of the physiological role of PBS movement in cyanobacteria.     

DnaJ-like protein gene sll1384 is involved in phototaxis in Synechocystis sp. PCC 6803

In Synechocystis sp. PCC 6803, gene sll1384 encodes a protein with a DnaJ domain at its N-terminal portion and a TPR domain at the C-terminal portion. An sll1384 mutant shows no difference from the wild type in adaptation to different temperatures, but almost completely loses its capability of phototactic movement. After complementation with sll1384, the mutant regains the phototaxis. As shown with electron microscopy, on the cell surface, mutant cells have pill that appear to be the same as that of the wild type. Also, the transformation efficiency remains unchanged in the mutant. It is postulated that Sll1384 regulates phototaxis of Synechocystis through protein-protein interaction. It is the first DnaJ-like protein gene identified in a cyanobacterium for a role in phototaxis.     

Role of two phosphohexomutase genes in glycogen synthesis in Synechocystis sp. PCC6803

Phosphohexomutases catalyze the interconversion between hexose-6-phosphate and hexose-l-phosphate and play important roles in polysaccharide synthesis. In Synechocystis sp. PCC 6803, sl10726 is predicted to encode PGM （phosphoglucomutase）, slr1334 is predicted to encode a PGM/PMM （phosphomannomutase） bifunction enzyme. In comparison to the wild type, a sllO726-null mutant showed 3.4% PGM activity but 45%-69% glycogen content. Down-regulation of slr1334, an essential gene, by using a copper regulated promoter further decreased the PGM activity in the sllO726：：Kmr PpetE-slr1334 double mutant to 0.3% of the wild type level. However, the glycogen content was not further decreased in parallel. In vitro, recombinant Sl10726 or S1r1334 showed predicted enzyme activities. Our results indicate that a relatively high level of glycogen can be maintained in Synechocystis mutants with low levels of PGM activity. The high PGM activity in the cyanobacterium may be required for turnover of glycogen or synthesis of other polysaccharides or oligosaccharides.     

all 1691基因表达及铁元素对鱼腥藻生长的影响

为实现FurA的表达,探究铁元素对藻生长的影响,首先运用Primer5.0设计引物,克隆出鱼腥藻PCC7120染色体上all1691(furA)基因片段;构建含组氨酸融合表达标签和T7启动子标签的表达载体.IPTG诱导FurA蛋白表达.然后设计不同Fe3+浓度梯度培养基,利用SDS-PAGE检测高、中、低Fe3+浓度培养液的藻细胞总蛋白,考马斯亮蓝g-250法测定蛋白含量,并用Trizol法提取不同Fe3+浓度培养的藻细胞总RNA.结果表明,获得纯化的all 1691克隆片段,大小为456bp,pET-1691重组蛋白在1mmol/L的IPTG诱导下,于37℃下摇床培养15h得到成功表达;低浓度Fe3+促进藻生长,高浓度Fe3+抑制藻生长;Fe3+浓度为2.0mg/L时rRNA位置与亮度清晰可见,而缺铁培养的藻生长几乎停滞,转录和翻译水平都很低.     

口服转胸腺素α1基因聚球藻对小鼠T细胞亚群作用的研究

为探讨口服后转胸腺α1(Tα1)基因聚球藻对T细胞亚群的作用,将小鼠随机分为空白对照组、野生藻组、转化藻(小剂量、中剂量和大剂量)组和日达仙组,用灌服的方式,给予空白对照组20mL·kg~(-1)的生理盐水,野生藻组0.4g·kg~(-1)的野生聚球藻;转化藻小、中、大剂量组,分别0.2g·kg~(-1)、0.4g·kg~(-1)和0.6g·kg~(-1)的转Tα1基因聚球藻;日达仙组则肌注给予3.2mg·kg~(-1)的胸腺素α1(日达仙)。结果表明转Tα1基因藻口服后可显著提高CD3亚群水平,显著提高CD4亚群水平,明显提高CD4/CD8的比值。提示转Tα1基因聚球藻具有增强机体免疫功能的作用。     

Effect of Iron Deficiency on Heterocyst Differentiation and Physiology of the Filamentous Cyanobacterium Anabaena sp. PCC 7120

0　IntroductionIron playsimportantrolesinthemetabolismofcyanobacteria,andisin volvedinavarietyofbiologicalactivities,bothasacofactorandcatalyst.Thesein cludephotosynthesis,pigmentsynthesisandfunction,respiration ,nitrateandnitritere duction ,hydrogenasereactions,dinitrogenfixation,andotherbiologicaloxidations.Numerousstudieshavebeencarriedoutontheeffectofnutritionaldeficiencyoncyanobacteria .Thephysiologicalandmolec ulareffectsofirondeficiencyoncyanobacte riahavebeenstudiedfromavarietyofper sp…     

鱼腥藻PCC 7120染色体上relBE同源基因的克隆及表达

为研究鱼腥藻PCC7120染色体上与relBE同源的基因asl4561和asl4562表达产物的毒素-抗毒素作用,从鱼腥藻细胞中提取总基因组DNA,设计特异引物PCR扩增目的基因asl4561和asl4562,与T载体连接后经XhoI和EcoR I双酶切并与表达载体pET28a（＋）连接,由IPTG诱导表达,并在IPTG浓度和诱导时间上对asl4561基因的表达条件进行了优化.琼脂糖凝胶电泳显示扩增出了264bp的asl4561基因和213bp的asl4562基因,SDS-PAGE检测表明成功表达出15.65kD和13.55kD的两蛋白,当诱导温度28℃、IPTG浓度0.4 mmol/L、诱导时间6h时,asl4561基因表达蛋白在细菌裂解液上清中表达量最大.     

Cloning and Characterization of the fecC Gene Necessary for Optimal Growth under Iron-Deficiency Conditions in the Cyanobacterium Anabaena sp. PCC7120

0　IntroductionIronisthefourthmostabundantelementbyweightintheearth’scrust.Itisrequiredfornearlyallorganisms.Itinvolvedinawidevarietyofbiochemicalprocessesbothasacofactorandcatalyst.Theseprocessesincludephotosynthesis,chlorophyllproduction ,respiration,nitrateandnitritereduction,nitrogenfixation ,andotherbi ologicaloxidations.However,itisanelementwithalowbiologicalavailabilityduetoitspoorsolubilityinoxygenicaqueoussolution .Livingor ganismsareconstantlyfacedtotheproblemofhowtoovercomethelows…     

磺基水杨酸的分子内质子转移及其在酸性锡电镀液中的荧光快速测定

根据磺基水杨酸反常大的荧光Ｓｔｏｋｅｓ位移的实验现象，提出了它在一定条件下发生分子内质子转移的机理．据此建立了酸性锡电镀液中磺基水杨酸的直接快速荧光分析新方法．该方法不受共存组分干扰，５～１０ｍｉｎ可完成测定，测定的相对标准偏差小于３％．     

鱼腥藻铁调蛋白基因(alr0957)表达和Fe~(3+)对藻生长的影响

根据CyanoBase提供的鱼腥藻PCC7120 FurC基因(alr0957)序列信息设计特异性引物,用降落PCR法从染色体DNA中扩增得约450 bp目的片段.运用TA克隆将其连接到pMD18-T载体上,经筛选测序获得阳性重组质粒.再经过双酶切、纯化FurC基因,连接原核表达载体pET-28a(+),并转化表达菌株BL21和IPTG诱导表达.经测序鉴定的阳性菌株中的FurC,运用SDS-PAGE检测重组蛋白,并利用镍柱层析法纯化目的蛋白.由藻细胞密度、叶绿素a含量、可溶性糖含量等改变探讨不同Fe3+浓度对藻类生长的影响.结果表明:在37℃经1 mmol/L IPTG诱导18 h,成功表达了分子量约为19000的融合蛋白.小于0.5mg/L Fe3+促进藻生长,大于0.9 mg/L Fe3+抑制藻生长,最适藻生长Fe3+浓度为0.5⁰.9 mg/L.     

Effect of iron deficiency on heterocyst differentiation and physiology of the filamentous cyanobacterium<Emphasis Type="Italic">Anabaena</Emphasis> sp. PCC 7120

The effect of iron deficiency on heterocyst differentiation and some physiological properties of the filamentous cyanobacteriumAnabaena sp. PCC 7120 was investigated. Under moderate iron limitation conditions, achieved by addition of iron chelator 2, 2′-Dipyridyl (<80 μmol/L) led to delayed heterocyst differentiation, no heterocyst differentiation was observed under severe iron limitation conditions, when the concentration of 2,2′-Dipyridyl in the medium was more than 100 μmol/L. it seemed that there are certain iron-regulated genes or operons whose function is to control heterocyst development. In addition, iron deficiency impaired the growth. Low iron cells had a decrease in the quantities of pigment content (chlorophyll and phycocyanin content), the whole cellin vivo absorbance spectra confirmed the decrease, the protein electrophoretic profiles revealed that iron-deficient cells had less protein bands, with the increase of 2,2′-Dipyridyl, the protein bands was more and more less. And differently, iron deficiency also caused an increase of ROS (Reactive Oxygen Species) and SOD activity, it suggests that iron deficiency led to oxidative stress, which generally occured under high-iron conditions. Foundation item: Supported by the National Natural Sciences Foundation of China (30070154), the Frontier Science Projects Programme of the Institute of Hydrobiology, the Chinese Academy of Sciences (220316), State Key Project on Cyanobacterial Bloom Control in Lake Danchi (K9905-35-01) Biography: Xu Wenllang (1974-), male, Ph. D. research direction: molecular genetics of eyanobacteria.     

溴氰菊酯降解菌Pseudomonas sp.P1-1-B3产酶条件的优化

农药降解酶对去除农药残留污染具有良好的应用前景.从海洋沉积物中筛选到一株高产溴氰菊酯降解酶的菌株Pseudomonas sp.P1-1-B3,研究了碳源、氮源、pH值、培养温度、培养时间、接种量及溴氰菊酯对菌株产酶的影响.结果表明,降解菌产酶的最适条件为:以可溶性淀粉作为碳源,m(蛋白胨):m(酵母浸膏)=2:1为氮源,pH 7.5,温度30℃,培养时间2 d,接种量3%,溴氰菊酯含量15μmol/L.在此条件下,菌株产酶的比活力最大为113.3 U/mg.该降解酶对溴氰菊酯的降解,在12 h内降解率达54.6%.