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1.
Partial rDNA sequences of Prorocentrum minimum and Takayama pulchella were amplified,cloned and sequenced,and these sequence data were deposited in the GenBank.Eight oligonucleotide probes(DNA probes)were designed based on the sequence analysis.The probes were employed to detect and identify P.minimum and T. pulchella in unialgal and mixed algal samples with a fuorescence in situ hybridization method using flow cytometry.Epifluorescence micrographs showed that these specific probes labeled with fluorescein isothiocyanate entered the algal cells and bound to target sequences,and the fluorescence signal resulting from whole-cell hybridization varied from probe to probe.These DNA probes and the hybridization protocol we developed were specific and effective for P.minimum and T. pulchella,without any specific binding to other algal species.The hyrbridization efficiency of difierent probes specific to P.minimum was in the order:PMl8S02＞PM28S02＞PM28S01＞PM18S01,and that of the probes specific to T. pulchella was TP18S02＞TP28S01＞TP28S02＞TP18S01.The djfferent hybridization efficiency of the DNA probes could also be shown in the fuorescent signals between the labeled and unlabeled cells demonstrated using flow cytometry.The DNA probes PM18S02,PM28S02,TPl8S02 and TP28S01,and the protocol,were also useful for the detection of algae in natural samples. 相似文献
2.
Partial rDNA sequences of Prorocentrum minimum and Takayama pulchella were amplified,cloned and sequenced,and these sequence data were deposited in the GenBank.Eight oligonucleotide probes(DNA probes)were designed based on the sequence analysis.The probes were employed to detect and identify P.minimum and T. pulchella in unialgal and mixed algal samples with a fuorescence in situ hybridization method using flow cytometry.Epifluorescence micrographs showed that these specific probes labeled with fluorescein isothiocyanate entered the algal cells and bound to target sequences,and the fluorescence signal resulting from whole-cell hybridization varied from probe to probe.These DNA probes and the hybridization protocol we developed were specific and effective for P.minimum and T. pulchella,without any specific binding to other algal species.The hyrbridization efficiency of difierent probes specific to P.minimum was in the order:PMl8S02＞PM28S02＞PM28S01＞PM18S01,and that of the probes specific to T. pulchella was TP18S02＞TP28S01＞TP28S02＞TP18S01.The djfferent hybridization efficiency of the DNA probes could also be shown in the fuorescent signals between the labeled and unlabeled cells demonstrated using flow cytometry.The DNA probes PM18S02,PM28S02,TPl8S02 and TP28S01,and the protocol,were also useful for the detection of algae in natural samples. 相似文献
3.
用双特异探针技术定性定量分析微型原甲藻 总被引：5，自引：0，他引：5
针对微型原甲藻核糖体大亚基和小亚基RNA分别设计了大亚基探针(LSU probe)和小亚基探针(SSU probe),发展了对微型原甲藻进行定性和定量分析的双特异探针技术.分析数据表明针对微型原甲藻核糖体大亚基RNA的LSU probe能够特异地将微型原甲藻和其他7种微藻分开,而且检测信号的强弱在一定的范围内与细胞数呈线性关系;由于针对微型原甲藻核糖体小亚基的SSU probe探针由于与具齿原甲藻存在核酸序列一致性,该探针与具齿原甲藻有严重的交叉反应,但未发现与海洋原甲藻和其他藻的交叉反应.另外,研究还优化了破碎微型原甲藻细胞所需的超声时间以及获得探针的特异性所需的S1酶反应温度.本研究为实现对微型原甲藻海水样品的快速准确监测奠定了基础. 相似文献
4.
共栖于对虾体表的致病性纤毛虫Ⅰ.(纤毛动物门,缘毛目) 总被引：1，自引：0，他引：1
宋微波 《中国海洋大学学报(自然科学版)》1991,(3)
采用现代通行的蛋白银染色法及Foissner干银法对共栖在对虾体表的钟虫属11种致病性纤毛虫做了形态分类学研究。报道了5种国内新记录。对所涉种类的银线系及纤毛下器做了首次揭示与描述,并列表对诸种的分类特征进行了统计学界定。 相似文献
5.
6.
Masahiko Nishimura Kumiko Kita-Tsukamoto Kazuhiro Kogure Kouichi Ohwada Usio Simidu 《Journal of Oceanography》1993,49(1):51-56
Fluorescent oligonucleotide probes were used to detectVibrio cholerae directly under epifluorescence microscope. The probe which is complementary to the specific 16SrRNA sequence ofVibrio cholerae was labelled with Texas-Red, whereas the universal probe was labelled with Fluorescein. These probes allowed the distinctionVibrio cholerae from other eubacteria under the same microscopic field. In order to detect and enumerate specific bacteria in natural seawater, this method was combined with the direct viable count (DVC) technique. The combined method increased intracellular rRNA levels in the sample, and made it possible to detect the target bacteria with the specific gene probe. The applicability of this new method was confirmed both for the laboratory mixed culture system and natural seawater. 相似文献
7.
对7株赤潮原甲藻28S rDNA 5'端部分序列进行扩增、克隆和序列测定,并从GeneBank上获取14个原甲藻28S rDNA序列,用NJ法和ME法构建了原甲藻属的系统树,并对序列进行分析.结果表明,7株原甲藻28S rDNA扩增序列长度为950~958 bp,通过NJ法和ME法构建的系统树完全一致.大部分分离自不同海域的同种原甲藻的序列高度保守,而不同种间在序列高变区却有较大的差异.但来自南海海域的海洋原甲藻(Prorocentrum micans)与分离自其他海域的株系序列差异较大,甚至超出了有些种间的差异.由28S rDNA高变区获得的序列,有望成为浮游植物特异性分子探针设计的良好靶区域. 相似文献
8.
Variations in the concentration of redox sensitive elements combined with pyrite framboid size data documented from a Marcellus Formation (Middle Devonian) core recovered from southwestern Pennsylvania elucidate the redox, organic matter accumulation, and diagenetic history of these deposits in this region of the basin. Uranium and Mo enrichment and Fe/Al display sharp increases coincident with diminishing Th/U upward through the initial 3rd order trangressive systems tract (lower Union Springs Member). These data as well as abundant small (<6 μm) pyrite framboids record establishment of strongly reducing benthic conditions, perhaps related to the expansion of an oxygen minimum zone induced by increased surface productivity. Strongly anoxic, even euxinic, conditions were interrupted by episodes of dysoxia, perhaps seasonal or longer term. Overlying regressive systems tract (RST) deposits record modestly improved redox conditions, likely a reflection of a receding oxygen minimum zone as base level dropped. A subsequent 3rd order base level rise and renewed expansion of the oxygen minimum zone triggered by increased surface productivity resulted in the accumulation of the organic-rich lower Oatka Creek Member. Still, the mix of abundant small and subordinate large (>10 μm) framboids preserves the record of oxygen deficient to sulfidic bottom conditions frequently interrupted by episodes of (dys)oxia. Improving redox conditions through the overlying RST were accompanied by a two-fold increase in Al and consequent dilution of the organic matter flux and authigenic trace metal uptake at the sediment–water interface. The upper half of the Oatka Creek comprises a depositional sequence not obvious from core inspection or gamma-ray signature but revealed by Mo enrichment and Al concentration profiles. Diagenetic modification of the Marcellus includes several horizons of authigenic calcium carbonate concretions and marked Ba enrichment. Both features reflect the effects of non-steady state microbial diagenesis within a methane-rich sedimentary column. 相似文献
9.
本研究将交叉引物恒温扩增技术(cross priming amplification,CPA)与核酸试纸条相结合建立一种副溶血性弧菌(Vibrio parahaemolyticus)快速可视化检测方法。针对副溶血性弧菌特有的不耐热溶血素基因tlh的六个不同区域设计两对特异性引物和一对检测探针,通过优化反应条件确定了最佳反应体系。CPA-核酸试纸条方法对副溶血性弧菌的检测具有较强的特异性,对纯培养物的检测灵敏度达到58 cfu/m L,对污染牡蛎中副溶血性弧菌的检测灵敏度为5.2 cfu/g,比传统的PCR技术灵敏度提高了10倍,且具有较高的稳定性。交叉引物恒温扩增技术与核酸试纸条相结合的方法操作简便、特异性强、灵敏度高且能有效防止污染,可用于现场及基层单位副溶血性弧菌的快速检测。 相似文献
10.
CHEN Guofu WANG Quanfu ZHANG Chunyun ZHANG Baoyu WANG Guangce LU Douding XU Zhong YAN Peishen 《海洋学报(英文版)》2013,32(2):66-75
Harmful algal blooms recently have been under the spotlight throughout the world, because of their negative impact on the marine environment, aquaculture, fisheries as well as public health. The development of methods for rapid and precise identification and quantification of causative species is essential for the warning and monitoring of blooms, among which the techniques based on taxonomic probes are the most favored. In this study, two harmful algae, i.e., Prorocentrum minimum and Karenia mikimotoi were taken into consideration. The partial large subunit rDNA (D1-D2) of both species were firstly PCR-amplified, cloned and sequenced. The obtained sequences were then introduced to carry out alignment analysis for gene specific regions. Three respective candidate probes for each species were designed and used to screen the optimal probe by performing fluorescence in situ hybridization (FISH) tests. The results showed that the probes Pmin0443 and Kmik0602 displayed the best hybridization for P. minimum and K. mikimotoi, respectively. Both the specific (taxonomic) (Pmin0443 and Kmik0602) and the control probes (UniC0512 and UniR0499) were used for cross-reactivity tests with other microalgae in our laboratory. The probes Pmin0443 and Kmik0602 are specific and could be served as taxonomic probes introduced into the techniques targeting rRNA, such as FISH, sandwich hybridization, and DNA-microarray assay of P. minimum and K. mikimotoi in the future. Finally, FISH analyses with both probes were performed on the simulated field samples. The probes could hybridize exclusively with the target cells well, and no significant difference (p >0.05) was observed in the cell densities of the samples determined by FISH and light microscopy (LM). All suggest that the probes are specific and could be introduced into FISH for the monitoring of both harmful algae. 相似文献
11.
赤潮叉角藻18SrDNA和ITS区序列测定与分析 总被引：20，自引：0，他引：20
采用PCR及克隆测序的方法，对1998年引发渤海赤潮的叉角藻18SrRNAadldey DNAITS区（Internal Transcribed Spacer Regions)进行了序列测定与分析。并通过因特网从国际分子生物学数据库中获取甲藻另外15个种的18rDNA序列，以Tetrahymena corlissi作为类群，分别采用Neighbor-Joining和Fitch方法构建甲藻较为一致和可靠的进化树图，探讨具有高度多样性和在分类上争议较多的甲藻各类群之间的形态与分子进化关系。结果表明，Prorocentrum(有2个简单的壳板）出现得较早，而大多数多甲藻目（覆盖着多个壳板）、裸甲藻目（大多数不具壳板）和膝沟藻目的成员较晚出现。另外，对叉角藻ITS区的分析表明，ITS区为高变区，是良好的分子标记，可用于叉角藻快速鉴定的专一性核酸分子探针的研制。 相似文献
12.
对笛鲷属9种鱼的线粒体DNA16S rRNA基因片段进行了PCR扩增,扩增产物经纯化后测序,得到了长度为561bp的分析序列。结合GenBank中的另外9种笛鲷的16S rRNA同源序列计算得出A、T、G、C的含量平均为28.5%、22.1%、23.6%和25.8%,AT含量稍高于GC含量,18种笛鲷碱基组成差异不大。利用MEGA3.1软件对所得序列进行比对后检测到72个变异位点,其中包括简约信息位点44个,在变异位点中75.61%的碱基替换是由于发生了转换,转换/颠换平均为3.1︰1。计算了种间的遗传距离,结果表明,序列差异在0.0193-0.0680之间,其中勒氏笛鲷和金焰笛鲷、勒氏笛鲷和金带笛鲷的序列差异最小,而红鳍笛鲷和金焰笛鲷、红鳍笛鲷和画眉笛鲷的序列差异最大。选用高体四长棘鲷(Argyrops spinifer)为外群,利用NJ法构建了分子系统树,结果表明:本研究中的9种南海笛鲷与其它9种笛鲷进化关系较远;并且南海9种笛鲷相互之间仍然保持着着相对较远的遗传距离,遗传多样性较好。 相似文献
13.
尽管18S rRNA基因序列极其保守,但是在一些种类中仍然发现其存在序列多态性。本研究在线纹舌鳎18S rRNA基因中发现了3种差异显著的序列类型（Type A,B和C）,说明其为非协同进化,而不是严格遵循协同进化方式。基于以上三种类型序列GC含量、二级结构、最小自由能等差异,我们推测Type A和B可能是假基因类型。此外,在Type A类型中还发现了一段长达189 bp的重复序列。据我们所知,如此长的重复序列,这在硬骨鱼类核糖体基因中还是首次报道。通过比较分析产生重复序列常见的模型,我们认为不等交换模型最有可能解释该重复序列的形成。本研究结果不仅为不等交换模型提供了罕见的分子证据,而且为鱼类18S rRNA基因研究奠定了很好的基础。 相似文献
14.
S7核糖体蛋白基因片段序列变异与花鲈属鱼类系统发育研究 总被引：1，自引：0，他引：1
对中国和日本海域花鲈属的3个种:花鲈(Lateolabrax maculatus)、鲈鱼(L. japoni-cus)和高体鲈(L. latus)的S7核糖体蛋白基因片段序列进行了扩增和序列测定,分析比较了3个种23个个体间的序列差异。在456bp的S7核糖体蛋白基因片段中,3个高体鲈个体中出现了3种单倍型,种内个体间有2个碱基差异;7个花鲈个体中出现了7种单倍型,种内个体间有21个碱基差异;13个鲈鱼个体中出现了12种单倍型,种内个体间有22个碱基差异。结果表明,花鲈属S7核糖体蛋白基因片段序列具有丰富的遗传信息,在亲缘关系较近的物种间也存在显著的序列差异,基于S7核糖体蛋白基因片段序列的NJ和ME系统树经1000次重复抽样检验后得到相似结果,表明S7基因内含子2是适合于研究分子系统发育的分子标记。利用Modeltest选取最佳核苷酸替代模型构建的NJ树表明:花鲈属鱼类由高体鲈分化,花鲈与鲈鱼的亲缘关系很近,而与高体鲈的亲缘关系较远。 相似文献
15.
利用Vector NTI Advance 11软件分析了NCBI数据库中大菱鲆的12428条EST序列,筛出候选SNP位点;应用高分辨率熔解曲线(HRM)对大菱鲆不同性状群体进行基因分型。结果表明,522个重叠群中共筛出160多个候选SNP位点;设计56对引物用于实验,能扩增出目的片段的引物有37条,合45个位点,成功率为66.1%;设计探针,能与候选SNP位点杂交的探针有13条,杂交率为28.9%。本实验中能成功进行基因分型的位点共有8个,占杂交位点的61.5%,其中有6个为碱基替换型位点,即G-A和C-T各占3个,2个为碱基颠换型位点,即T-G和A-T各占1个。 相似文献
16.
An experimental investigation was conducted in the laboratory in order to evaluate the use of fiber-optic probes for measuring the size distribution of large transient bubbles entrained beneath breaking ocean waves. Measurements were made in a unidirectional flow cell using two-fiber Optoflow fiber-optic probes (Photonetics Inc.). It was found that the rise times of the signal pulses created when bubbles crossed the probe tip were closely correlated with the bubble velocities. Therefore, bubble velocities and hence bubble sizes could be estimated using a single probe. It was observed that the correlation between the rise time and the bubble velocity varied significantly between probes but was only a weak function of water type (i.e., freshwater or seawater) and the bubble impaction angle. Estimates of the bubble-size distributions and void fraction made using the fiber-optic probes and video image data were compared. It was found that the fiber-optic probes could be used to obtain fairly accurate estimates of the void fraction. The bubble-size distributions estimated from the probe signals compared favorably with those estimated from video images. It was concluded that the probes could be used to make reasonably accurate measurements of the bubble-size distributions beneath breaking ocean waves 相似文献
17.
赤潮异弯藻和海洋原甲藻LSU Rdna扩增及序列分析 总被引：1，自引：1，他引：1
利用引物D1R和D2C扩增并测定了赤潮异弯藻(Heterosigma akashiwo Hada)和海洋原甲藻(Prorocentrum micans Ehrenberg)的LSU rDNA D1与D2序列,并与GenBank中相关序列进行比较分析.结果表明:在种内水平,所测的H.akashiwo 6个株系之间共有5个变异位点,序列H.k与H.k-2,H.k-4均存在碱基替换;原甲藻属不同种内各株系之间的遗传距离为0.002～0.023之间,所测序列P.mi与P.micans其他株系之间均存在碱基替换.在种间水平上,原甲藻属不同种之间的遗传距离在0.045～0.139之间,大于种内遗传距离,每个种都具有特定的保守序列.根据序列间的遗传变异,可设计特异性的探针对不同株系和物种进行检测和计数. 相似文献
18.
Ya. A. Virolainen Yu. M. Timofeev D. V. Ionov A. V. Poberovskii A. M. Shalamyanskii 《Izvestiya Atmospheric and Oceanic Physics》2011,47(4):480-490
To interpret the ground-based measurements of the spectra of direct solar infrared radiation with the help of a Brucker Fourier-spectrometer, a technique for determining the total ozone content (TOC) was developed and implemented. The TOC was determined using six spectral intervals of an ozone-absorption band of 9.6 μm and the shortwave panel of a carbon-dioxide-absorption band of 15 μm, where the impact of other atmospheric parameters on the measured solar radiation was reduced to a minimum. The potential errors of the infrared method for determining the TOC for the chosen spectral scheme with the influence of measurement errors and vertical profiles of temperature are less than 1% for different signal-to-noise ratios and zenith angles of the sun. We analyzed 269 high-resolution (0.005–0.008 cm?1) spectra of solar infrared radiation measured in Peterhof over 52 days from March to November, 2009. The resulting values of TOC were compared with the results of independent ground-based TOC measurements in Voeikovo (Main Geophysical Observatory) using a Dobson spectrophotometer and an M-124 ozonometer, as well as with the Ozone Monitoring Instrument (OMI) satellite data. The mean errors between the results of TOC measurements with the help of the three ground-based probes constitute no more than 0.4%. The rms errors between data obtained by the Brucker spectrometer and the given satellite and ground-based probes constitute 3–4%. A comparison between different series of measurements indicated that the upper estimate for the error of TOC measurements by the Brucker spectrometer was 2.5–3% (when the possible spatial and temporal errors in measurements are disregarded). An analysis of the diurnal variations in the TOC measurements for stable atmospheric conditions yields an upper estimate of ～3 DU (around 1%) for the random component of error in TOC measurements by the Brucker spectrometer. 相似文献
19.
Bennett R. Burns J. Nastav F. Lipkin J. Percival C. 《Oceanic Engineering, IEEE Journal of》1985,10(1):17-22
Two single-sensor piezometer probes, 8 mm in diameter, were developed for deep-ocean geotechnical investigations. These probes were tested in a hyperbaric chamber pressurized to 55 MPa (8000 psi). Testing was performed for a period of five weeks under high hydrostatic pressure with the probes inserted in reconstituted illitic marine sediment. Small differential pore-water pressures were generated in response to both mechanically and thermally generated forcing functions. During deep-ocean simulated pressure tests, the sensors exhibited excellent sensitivity and stability. These developments in piezometer-probe technology provide a quantitative means of assessing important geotechnical parameters of fine-grained seabed deposits. 相似文献
20.
M. Paul Mosley 《新西兰海洋与淡水研究杂志》2013,47(3-4):351-357
Riffles and points of divergence of branch channels in braided rivers are critical controls upon passage of fish and recreational boats because they are points at which depths are at a minimum. Depths on randomly selected riffles were measured in braided reaches of the Ashley, Hurunui, and Rakaia Rivers and related to discharge; minimum depths encountered in extended reaches of these rivers at a range of flows have also been measured. The data can be used to predict minimum available passage depths at a specified flow, or conversely, to predict the discharge required to maintain a specified minimum passage depth. However, presently available estimates of critical minimum depths required for various instream uses (migration of salmonids, jetboating) appear excessively conservative, and minimum depth requirements must be more accurately determined before instream flow needs can be properly assessed. 相似文献