The research of a high-cell density cultivation of Synechococcus sp.PCC7942, transformed of UB-Tα1

蓝藻（cyanophyte），又称蓝细菌（cyanobacteria），是一类光合自养型原核生物。多数蓝藻富含营养物质、无毒，常作为外源目的基因的表达系统而被广泛应用于分子生物学领域的研究。 本实验室利用以蓝藻为受体的穿梭质粒表达载体和基因整合平台打靶载体两套系统，将人源胸腺素α1基因转入到蓝藻Synechococcussp.PCC7942表达载体中，筛选出高效表达人源胸腺素α1的转基因藻株，并检测到活性胸腺肽α1的表达。 在成功地构建转基因藻之后，实现高密度培养和高效表达外源目的基因是转基因藻能否走向应用的关键环节之一。本研究以蓝藻无机培养基BG-11为基础，探讨接种量和各种理化因子如，...Cyanobacteria, or cyanophyte are photoautotrophic organisms with many special properties, making them very attractive for molecular biological studies. Funded by National High Science & Technical Program (863), our lab had succed in expressing human thymosinα1 (Tα1) gene in cyanobacteria Synechococcus sp.PCC7942 with the suage of shutter vector system and reconstructed plasmid vector system. Thi...学位：理学硕士院系专业：生命科学学院生物学系_微生物学学号：19992600

Zhongshan Wang’s calligraphic and painting hanging scroll - 汪仲山字與畫

Kun Wang, known as Zhongshan (courtesy name), was born in Jiangxi, China. Wang is a renowned artist specializing in landscape and flower painting at the beginning of Republic China. His style was influenced by Hui Wang (1632-1717, a Chinese landscape painter).汪琨，字仲山，江西人。擅長山水、花卉畫，畫風受王翬影響。1) Ancient saints have remarkable ambition. When they develop a virtue as God’s grace, there are usually four signs indicating their achievement. First, they show honesty, respect for other people and extraordinary self-cultivated knowledge. These are also qualities stressed by Yi Cheng (1033-1107, a Chinese philosopher during Song dynasty). Second, they are so sincere. When they pray, God moved and gave them ability of foresight. This is also a personality that Zisi recommended (483-402 B.C., a Chinese philosopher and only grandson of Confucius). Third, they content with a simple life and show loyal to their existing belief. Such attitude benefits their spiritual health and makes them shine and look rosy about the gills. These characteristics are also critical features addressed by Confucius (551-479 BC, a Chinese politician, philosopher and educator), Hui Yan, (521-490 BC, one of the disciples of Confucius), Zengzi (505-437 BC, a Chinese philosopher and student of Confucius) and Mencius (a Chinese philosopher). Fourth, they are able to see true nature of objects and sing as they please. This ability leads to leisurely and placid sentiment and joyful spirits. It is also a skill pursued by Qian Tao (372?-427, a Chinese poet in Jin dynasty), Juyi Bai (772-846, a Chinese poet in Tang dynasty), Shi Su (1037-1101, a poet of Song dynasty) and You Lu (1125-1210, a Chinese poet in Southern Song dynasty). I regret that I did not work hard enough in my youth and now, at my old age, I fear and feel sorry for not realizing a state of mind of ancient saints. After I contemplate back and forth, I keep sighing for my deficiency. 2) In a sunset landscape, a river meanders to a lonely city at the horizon. Returning swallows is flying at the edge of the sky, passing mists around trees. I was shocked that winds moving between bamboos are so brutal. Cold airs are kept blown into houses through a thatch door.1) 古來聖哲胸懷極廣，而可達於德者，約有四端：如篤恭修己而生睿智，程子之說也；至誠感神而致前知，子思之訓也；安貧樂道而潤身睟面，孔彥曾孟之旨也；觀物閒吟而意適神恬，陶白蘇陸之趣也。自恨少壯不知努力，老年常多悔懼，於古人心境，不能領取一二。反覆尋思，嘆喟無已。 2) 夕陽流水遠繞孤村，數點歸鴉煙樹昏。怪底竹風無賴甚，又吹寒風入柴門。Postscript: Emulating Mengquanwaishi’s (Gang Xi, 1746-1803) style; Signature: 1) [Illegible]; 2) Zhongshan, Kun Wang in June, 1925.題跋: 臨蒙泉外史筆; 款識:1) [未辨識][未辨識]正辛未二月日記[未辨識][未辨識]; 2) 乙丑六月仲山汪琨1) An emperor-giving phase: keeping examining the past while facing challenges; 2) Ci-shu Tang; 3) Tse Tsung Chow; 4) Blessing-and-longevity cottage; 5) Kun Wang; 6) Zhongshan1) 御賜視履考祥; 2) 賜書堂; 3) 周策縱; 4) 福壽廬; 5) 汪琨; 6) 仲

High-cell density cultivation of transgenic Synechococcus sp. PCC7942 with human thymosin α_l gene

应用响应面分析方法(Response Surface Method,RSM),对转人源胸腺肽α1基因聚球藻7942的培养基及培养条件进行优化,优化后摇瓶培养转化藻的比增殖速率(μ值)可达0.500d-1,10 L全自动光生物反应器中培养转化藻μ值可达0.979 d-1,分别比正交实验提高了33%和161.1%,建立高密度培养的数字模型预测实验结果,预测值与实验值之间的相对误差在±5.0%以内,为响应面方法在微藻细胞培养中的广泛应用奠定了基础。The response surface method(RSM)was used to optimize the medium and culture conditions for transgenic Synechococcus sp.PCC7942 with human thymosin α1 gene.The results were as follows: NaHCO3 36 mg/L,KNO3 1.3 g/L,initial pH 10.0,light intensity 1800 lx.Under these conditions,the specific growth rate(μ) of the transgenic strain could reach 0.500 d-1 through shake-flask culture,when cultivated in 10L autophotobioreactor,the μcould increase to 0.979 d-1,with the maximal yield of 0.299 g/ L·d.There was no reduction of the bioactivity of UB-Tα1 in these culture processes

Construction of Intergeneric Yeast Fusant for ProductionSorbitol from Jerusalem Artichoke

介绍高产菊粉酶的克鲁维酵母 (K luyveromyces sp.) Y85菌株与产山梨醇的酿酒酵母(Saccharomyces cerevisiae) E15菌株 ,通过属间原生质体融合构建直接利用菊粉产山梨醇的酵母融合菌株 .结果认为 ,以 0 .1% EDTA-巯基乙醇为脱壁预处理剂 ,蜗牛酶浓度 2 % (W/V) ,酶解温度 30℃ ,E15和 Y85两菌株细胞酶解时间分别为 4 0 min和 30 min,原生质体形成率和再生率分别达90 %和 2 5 %以上 .融合的适宜条件为 :添加 30 % (W/V) PEG和 10 mmol/L Ca Cl2 ,融合时间为 30min,融合率可达 2 .64× 10 -6 以上 .利用两亲株自然性状差异检出融合菌株 ,并经 DNA含量、菊粉酶活性、细胞形态与大小、菌落形态等测定加以确证 .其中 F2 7融合菌株在适宜条件下 ,山梨醇摇瓶发酵产量可达 4 .87g/10 0 m L,并且具有遗传稳定性能The construction of inulin utilizing and sorbitol producing fusants by interge neric protoplast fusion between Kluyveromyces sp. Y 85 and Saccharomy ces cerevisiae E 15 is discussed. The strains protoplast were prepared with 0.1 % EDTA and 2.0%(w/v)helicase at 30 ℃ for 30~40 min, The formation rate was above 9 0% and regeneration rate reached 25%. The suitable conditions for the protoplas t fusion was added with 10mmol/L each and 30%PEG(MW6,000)for 30 min. The fusio n frequency was about 2.64×10 -6 . The fusants were sorted by using the d ifferent character between strans Y 85 and E 15. DNA estimation, inulinase ac tivity examination, and comparison of cell size and morphology indicated that the fusants were fused from strans E 15 and Y 85. Under optimal fermentation c onditions, a maximum sorbitol production reached 4.87g/100mL, and the fermenta tion ability of F27 fusant was quite stable

用硅离子注入SiO_2层方法制备的纳米硅的光学性质

报道了用硅离子注入热氧化生长的SiO_2层后热退火的方法制备纳米硅样品,并在室温下测量了样品的光致发光谱及其退火温度的关系。实验结果表明,在800℃以下退火的样品的发光是由于离子注入而引入SiO_2层的缺陷发光,在900℃以上退火,才观察到纳米硅的发光,在1100℃下退火,纳米硅发光达到最强,纳米硅的发光峰随退火温度升高而红移呈量子尺寸效应。在直角散射配置下,首次观察到纳米硅的特征拉曼散射峰,进一步证实了光致发光谱的结果

Optimization of culture conditions for transgenic Synechococcus sp. PCC7942 and wild-type strain

通过考察培养温度、光照强度、培养基初始pH和碳、氮源浓度等单因子对转人源胸腺肽α1基因聚球藻PCC7942及其野生藻生长的影响,对其中NaHCO3浓度、KNO3浓度和初始pH进行三因素三水平的正交试验,获得以BG-11为基础的培养基成分的最佳组合及最适培养条件.实验结果表明,转化藻于28℃,光照强度1500Lx,往复式光照摇床转速75r/min,初始pH=9.0,NaH-CO330mg/L,KNO31.7g/L(其他同BG-11培养基)培养;野生藻于28℃,光照强度2000Lx,往复式光照摇床转速75r/min,初始pH=9.0,NaHCO340mg/L,KNO32.2g/L(其他同BG-11培养基)培养,藻细胞生物量均得到有效地提高,其比增殖速率分别达0.375d-1和0.382d-1,比优化前分别增加106%和99%,这为微藻的进一步高密度放大培养提供依据. The requirements of carbon source, nitrogen source, light intensity, initial pH and temperature by transgenic Synechococcus sp. PCC7942 with human thymosin α_1 gene and its wild-type strain were optimized by means of orthogonal tests based on the result of single factor experiments. The optimal cultural conditions of the transgenic strain based on BG-11 medium were determined as follows: 28 ℃, initial pH 9.0, light intensity 1 500 Lx, rotating speed 75 rpm, NaHCO_3 30 mg/L, KNO_3 1.7 g/L(the rest of the components were the same as BG-11 medium), as well as light intensity 2 000 Lx, NaHCO_3 40 mg/L, KNO_3 2.2 g/L of the wild-type strain, which evidently increased the cell biomass of these two cyanobacteria. The results showed that under optimal conditions the specific growth rate of the wild-type strain and the transgenic one reached 0.382 d~(-1) and 0.375 d~(-1) respectively, which were 99% and 106% higher than the unoptimization.国家海洋863基金资助项目(819-04-03

Cultivation of transgenic Synechococcus sp. PCC7942 with human thymosin α_1 gene and wild-type strain

探讨光照强度、光照时间、接种量和不同碳氮源对转人源胸腺肽α1基因聚球藻 794 2 (转化藻 )及其野生藻 (聚球藻 794 2 )生长的影响以及生长过程中pH的变化。研究发现 ,无论是野生藻还是转化藻的生长都表现出全天光照优于半天光照。三角瓶培养转化藻生长的最佳光照度为15 0 0lx ,野生藻生长的最佳光照度为 2 0 0 0lx ;光反应器培养两种藻生长的适宜光照强度均随藻体浓度的变化而改变。为了尽量缩短延滞期 ,转化藻接种量OD730 ≥ 0 .0 8,野生藻OD730 ≥ 0 .0 5。两种藻体生长适宜的碳氮源为NaHCO3 和KNO3,生长过程中pH值变化趋势基本相似。Effects of light intensity, inoculum design and different medium components on the culture of transgenic Synechococcus sp.PCC7942 with human Thymosin α 1 gene and wild-type strain were studied. Both of the strains grew better in whole-day light. In flask, the optimal light intensity for transgenic strain was 1500lx, while that for wild-type was 2000lx. But the optimal light intensity of both strains was changed in airlift photobioreactor. The changes were relied on cell concentration during culture. The lay time was greatly reduced for transgenic strain when the inoculum cell concentration was more than 0.08 of OD 730, while for wild-type strain it was more than 0.05 of OD 730. NaHCO 3 and KNO 3 were found to be the best carbon and nitrogen source. The chose pattern of pH was almost the same for the two stains.国家海洋 86 3项目 (项目号 819-0 4-0 3)部分研究工

Optimum conditions for sorbital fermeration by a yeast fusion F27

报道了酵母属间融合菌株F2 7利用菊粉进行山梨醇发酵适宜条件的研究结果。其适宜的发酵培养基组成为 (% ) :菊粉 (以总糖计 ) ,10 ;酵母膏 ,2 .5 ;葡萄糖 ,1.0 ;MnSO4 ,0 .1。适宜的发酵条件为 :培养基初始 pH5～ 6 ,接种量 10 % (V/V) ,温度 35℃。该菌株经摇瓶发酵 72h后 ,山梨醇产量可达 4 .87g/ 10 0mLThe optimum conditions for sorbitol production from Jerusalem artichoke by a yeast fusant F27 were investigated. The suitable medium for the sorbitol production was composed of 10% inlulin(total sugar), juice, 2.5%yeast extract and 1.0% glucose. The optimum cultivation temperature and initial pH for the sorbitol production were 35℃ and 5～6 respectively. Under these conditions, a maximum sorbitol production of 4.87 g/100mL was obtained after 72 hours of growth in shake cultures