铜绿微囊藻Microcystis aeruginosa PCC7820 ATP含量和细胞分裂昼夜节律的研究

为了研究铜绿微囊藻Microcystis aeruginosa PCC7820中是否存在内源昼夜节律,检测了其细胞中的三磷酸腺苷(ATP)含量和细胞分裂的日变化规律.通过生物发光法检测的细胞内ATP含量变化结果表明,在12h光/12h暗(L/D)周期环境下该藻的ATP含量呈现明显的近似24h的昼夜周期性变化,且这种周期性变化在连续光照条件下能至少持续运行3个周期,周期长度具有温度补偿效应, 而光照和温度的改变能重置昼夜节律的时相,这说明铜绿微囊藻ATP含量的昼夜节律变化受到内源生物钟的控制.通过对细胞数、细胞大小和细胞分裂素含量的检测发现,传代时间为38.4h的铜绿微囊藻的细胞分裂也呈现受生物钟调控的昼夜节律性,这种昼夜节律变化可能是生物钟通过一种门控机制调控M. aeruginosa PCC7820的细胞分裂时相而产生的.     

铜绿微囊藻Microcystis aeruginosa PCC7820生物钟基因的克隆及其蛋白相互作用分析

为了深入研究铜绿微囊藻生物钟调控的分子机制,根据蓝藻钟基因的同源序列设计合成了简并引物,通过PCR和染色体步行的方法获得了铜绿微藻Microcystis aeruginosa PCC7820的生物钟基因簇kaiABC.kaiA基因全长873bp,编码一个由290αα组成,分子量大小为33.3kDa的蛋白;kaiB基因全长315bp,编码分子量大小为11.8kDa的蛋白;kaiC基因全长1563 bp,编码蛋白分子量大小为58.3kDa.Kai蛋白间的相互作用是昼夜节律计时产生的重要过程,利用酵母双杂交系统检测了三种Kai蛋白间的相互作用,结果表明KaiA、KaiB和KaiC蛋白自身及KaiA与KaiC、KaiB与KaiC之间都能发生明显的相互作用,而KaiB和KaiA之间没有检测到相互作用.     

铜绿微囊藻生物钟蛋白的表达纯化与抗体制备

为了获得融合表达的铜绿微囊藻(Microcystic aeruginosa)生物钟蛋白KaiA、KaiB、KaiC并制备其相应的多克隆抗体,将kaiA、kaiB、kaiC基因分别克隆到原核表达质粒pET-His中.重组质粒pET-His-KaiA,pET-His-KaiB和pET-His-KaiC经酶切和测序鉴定后,分别转化E.coli BL21(DE3)进行融合表达.经SDS-PAGE分析可知,融合表达的KaiA、KaiB和KaiC蛋白表达量可分别达到菌体总蛋白的25%、40%和20%.经亲和层析后融合蛋白KaiA和KaiB的纯度分别达95%和92%,而KaiC经胶回收纯化后纯度也可达93%.将纯化后的三种Kai蛋白作为抗原分别免疫小鼠制备多克隆抗体,经ELISA检测抗体滴度表明,制备的抗KaiA、抗KaiB和抗KaiC的多克隆抗体效价高,分别可达到1:50000、1:60000和1:100000.Western blotting结果表明:获得的多克隆抗体具有较高的效价,抗体能识别相应的Kai蛋白,具有较高的特异性,能用于铜绿微囊藻生物钟蛋白KaiA、KaiB和KaiC的表达节律检测.     

铜绿微囊藻(Microcystis aeruginosa)蛋白质组学研究方法的初步建立

参照一些植物蛋白的提取方法,初步确定了铜绿微囊藻总蛋白的提取方法,即TCA-丙酮法,并采用蛋白质组学技术路线,即双向凝胶电泳(2-D PAGE)及基质辅助激光解吸离子化-飞行时间质谱(MALDI-TOF)分析,建立了铜绿微囊藻在对数生长期的蛋白质组分子解剖图谱.将铜绿微囊藻接种于BG-11培养基28℃培养至对数生长期,提取藻种总蛋白进行双向电泳.从2-D PAGE图谱中随机选取110个蛋白质点进行肽质量指纹图谱(PMF)鉴定,检索发现21个确定为数据库中收录的蓝藻的基因或蛋白,其中10个为数据库中收录的聚球藻Synechococcus sp.PCC6803或微囊藻的基因或蛋白.     

微囊藻细胞计数法研究

本文阐述了用超声波短时作用来处理水样,从而找到更科学地计数水体中微囊藻细胞数的方法。并通过实验进一步研究了超声波频率和作用时间对处理效果的影响。     

微囊藻毒素分析检测技术研究进展

微囊藻毒素是一种在蓝藻水华污染中出现频率最高、产毒量大、危害严重的一类藻类毒素。微囊藻毒素不仅可以在动物体内蓄积,还可通过水产品进入人体,从而对人类健康造成潜在的威胁,本文综述了微囊藻毒素分析检测方法,并对其优缺点加以比较。     

饮用水中微囊藻毒素污染控制技术研究进展

随着饮用水源水污染的不断加剧,常规饮用水净水工艺已很难使出水水质达到国家新的饮用水卫生标准。因此,寻求经济、有效的处理技术,以保障饮用水的安全性,成为学科领域内研究热点。     

光合作用与应用

生物体内光合作用很多,能转化大气中过多的二氧化碳转化为氧气,制造有机物。我们通过研究光合作用将有助于我们设计出人造自然光捕捉系统,设计出入造光源能量转换装置,解决能源匮乏。毫不夸张地说,这项研究为我们开启了一扇通往光明的大门。在不远的将来,可以利用人造合成装置,在吸收大气中二氧化碳的同时,产生出入们所需要的能量,在实现环保的同时还能解决能源的制造问题。     

福州山仔水库富营养化和微囊藻毒素污染水平初步调查分析

目的:调查福州山仔水库的富营养化状况和微囊藻毒素污染水平。方法:采用固相萃取-高效液相色谱法对山仔水库水源地的微囊藻毒素进行为期一年的监测,同时测定与水体富营养化相关的水质指标。结果:表明2007年山仔水库在春末和夏、秋季检出了3种微囊藻毒(MC-YR、MC-RR、MC-LR),以MC-YR为主。山仔水库水体的营养状况为中营养到富营养化水平。夏季是藻毒素污染的高峰时段,其浓度与水库环境条件及水质的营养水平有密切的关系。     

模拟光合作用的新型太阳能电池问世

<正>据媒体报道,一种模仿大自然光合作用的新型太阳能电池日前问世,它可以利用太阳光能将无机物转化为有机物并贮存能量,并且能应用到多种消费品中。     

Tuning the phase of circadian entrainment

The circadian clock coordinates daily physiological, metabolic and behavioural rhythms. These endogenous oscillations are synchronized with external cues (‘zeitgebers’), such as daily light and temperature cycles. When the circadian clock is entrained by a zeitgeber, the phase difference ψ between the phase of a clock-controlled rhythm and the phase of the zeitgeber is of fundamental importance for the fitness of the organism. The phase of entrainment ψ depends on the mismatch between the intrinsic period τ and the zeitgeber period T and on the ratio of the zeitgeber strength to oscillator amplitude. Motivated by the intriguing complexity of empirical data and by our own experiments on temperature entrainment of mouse suprachiasmatic nucleus (SCN) slices, we present a theory on how clock and zeitgeber properties determine the phase of entrainment. The wide applicability of the theory is demonstrated using mathematical models of different complexity as well as by experimental data. Predictions of the theory are confirmed by published data on Neurospora crassa strains for different period mismatches τ − T and varying photoperiods. We apply a novel regression technique to analyse entrainment of SCN slices by temperature cycles. We find that mathematical models can explain not only the stable asymptotic phase of entrainment, but also transient phase dynamics. Our theory provides the potential to explore seasonal variations of circadian rhythms, jet lag and shift work in forthcoming studies.     

极大节旋藻(Arthrospira maxima)生物钟基因kaiC的克隆及分析

以kaiC基因簇部分已知序列为引物设计位点,采用PCR反应池法从节旋藻基因组fosmid文库中筛选到kaiC基因克隆,通过步移测序获得了kaiC基因全长序列。kaiC基因编码区长1554bo,基因GC碱基含量平均为41．76％,密码子第三位明显偏向于U。在基因上游385bp范围内发现一个可能的启动子序列和一些基因调控元件。KaiC蛋白分析发现了Walker A、DXXG、不完整的Walker B等重要模体和具有催化作用的功能位点。Southem杂交的结果证明kaiC基因在极大节旋藻中为单拷贝。极大节旋藻kaiC基因的特殊结构特征为研究kai基因簇的进化提供了重要的启示。     

Regulatory principles and experimental approaches to the circadian control of starch turnover

In many plants, starch is synthesized during the day and degraded during the night to avoid carbohydrate starvation in darkness. The circadian clock participates in a dynamic adjustment of starch turnover to changing environmental condition through unknown mechanisms. We used mathematical modelling to explore the possible scenarios for the control of starch turnover by the molecular components of the plant circadian clock. Several classes of plausible models were capable of describing the starch dynamics observed in a range of clock mutant plants and light conditions, including discriminating circadian protocols. Three example models of these classes are studied in detail, differing in several important ways. First, the clock components directly responsible for regulating starch degradation are different in each model. Second, the intermediate species in the pathway may play either an activating or inhibiting role on starch degradation. Third, the system may include a light-dependent interaction between the clock and downstream processes. Finally, the clock may be involved in the regulation of starch synthesis. We discuss the differences among the models’ predictions for diel starch profiles and the properties of the circadian regulators. These suggest additional experiments to elucidate the pathway structure, avoid confounding results and identify the molecular components involved.     

Modelling the functional roles of synaptic and extra-synaptic γ-aminobutyric acid receptor dynamics in circadian timekeeping

In the suprachiasmatic nucleus (SCN), γ-aminobutyric acid (GABA) is a primary neurotransmitter. GABA can signal through two types of GABA_A receptor subunits, often referred to as synaptic GABA_A (gamma subunit) and extra-synaptic GABA_A (delta subunit). To test the functional roles of these distinct GABA_A in regulating circadian rhythms, we developed a multicellular SCN model where we could separately compare the effects of manipulating GABA neurotransmitter or receptor dynamics. Our model predicted that blocking GABA signalling modestly increased synchrony among circadian cells, consistent with published SCN pharmacology. Conversely, the model predicted that lowering GABA_A receptor density reduced firing rate, circadian cell fraction, amplitude and synchrony among individual neurons. When we tested these predictions, we found that the knockdown of delta GABA_A reduced the amplitude and synchrony of clock gene expression among cells in SCN explants. The model further predicted that increasing gamma GABA_A densities could enhance synchrony, as opposed to increasing delta GABA_A densities. Overall, our model reveals how blocking GABA_A receptors can modestly increase synchrony, while increasing the relative density of gamma over delta subunits can dramatically increase synchrony. We hypothesize that increased gamma GABA_A density in the winter could underlie the tighter phase relationships among SCN cells.     

纳米材料在观赏鲤鱼养殖中的应用研究

试验通过室内箱体养殖鲤鱼,研究了纳米材料对鱼苗生长和对水体理化性状的影响。结果表明,纳米材料可提高水体pH值,降低NO2^-和NO3^-含量,改善水质;纳米材料处理水体可明显提高鱼苗的成活率,其中纳米网效果最好,使成活率提高一倍。     

An exploration of the utility of mathematical modeling predicting fatigue from sleep/wake history and circadian phase applied in accident analysis and prevention: the crash of Comair Flight 5191

On 27 August 2006 at 0606 eastern daylight time (EDT) at Bluegrass Airport in Lexington, KY (LEX), the flight crew of Comair Flight 5191 inadvertently attempted to take off from a general aviation runway too short for their aircraft. The aircraft crashed killing 49 of the 50 people on board. To better understand this accident and to aid in preventing similar accidents, we applied mathematical modeling predicting fatigue-related degradation in performance for the Air Traffic Controller on-duty at the time of the crash. To provide the necessary input to the model, we attempted to estimate circadian phase and sleep/wake histories for the Captain, First Officer, and Air Traffic Controller. We were able to estimate with confidence the circadian phase for each. We were able to estimate with confidence the sleep/wake history for the Air Traffic Controller, but unable to do this for the Captain and First Officer. Using the sleep/wake history estimates for the Air Traffic Controller as input, the mathematical modeling predicted moderate fatigue-related performance degradation at the time of the crash. This prediction was supported by the presence of what appeared to be fatigue-related behaviors in the Air Traffic Controller during the 30 min prior to and in the minutes after the crash. Our modeling results do not definitively establish fatigue in the Air Traffic Controller as a cause of the accident, rather they suggest that had he been less fatigued he might have detected Comair Flight 5191's lining up on the wrong runway. We were not able to perform a similar analysis for the Captain and First Officer because we were not able to estimate with confidence their sleep/wake histories. Our estimates of sleep/wake history and circadian rhythm phase for the Air Traffic Controller might generalize to other air traffic controllers and to flight crew operating in the early morning hours at LEX. Relative to other times of day, the modeling results suggest an elevated risk of fatigue-related error, incident, or accident in the early morning due to truncated sleep from the early start and adverse circadian phase from the time of day. This in turn suggests that fatigue mitigation targeted to early morning starts might reduce fatigue risk. In summary, this study suggests that mathematical models predicting performance from sleep/wake history and circadian phase are (1) useful in retrospective accident analysis provided reliable sleep/wake histories are available for the accident personnel and, (2) useful in prospective fatigue-risk identification, mitigation, and accident prevention.