921 resultados para AMPLIFICATION


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The aim of this study is to investigate the effects of different doses of 100 MeV/u carbon ions on sweet sorghum seeds in order to improve crop yields and their sugar content. After irradiation, seeds were germinated and grown to 30 days, and others were sown in the field. At the end of harvesting season all planted seeds were picked separately and M2 generations obtained. The differences among the treatments were examined using the RAPID procedure. In the study done by using 38 primers; according to the amplification results, the differences among the various doses treatment were shown.

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利用X射线对体外培养的人肝癌细胞SMMC-7721和正常人肝细胞HL-7702进行辐照,以细胞克隆形成法测定细胞存活率,多聚酶链式反应银染端粒重复序列扩增法(Telomeric repeat amplification protocol,TRAP-PCR)检测细胞中端粒酶活性变化,探讨辐照后细胞存活与细胞中端粒酶活性变化的关系。结果表明,SMMC-7721和HL-7702细胞的存活率都随辐照剂量的增加而相应降低。在1-4Gy剂量范围内,两种细胞的端粒酶活性变化均呈剂量依赖性增强。但在1Gy、2Gy、3Gy剂量下,SMMC-7721其端粒酶活性较对照(0Gy)细胞低;当辐照剂量达4Gy时,端粒酶活性略高于对照细胞。在1-4Gy剂量范围内,HL-7702和SMMC-7721的细胞存活与端粒酶活性在变化趋势上呈负的关系。

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端粒是染色体末端由重复DNA序列和相关蛋白组成的一种特殊结构,具有稳定染色体结构及完整性的功能,会随染色体复制与细胞分裂而缩短。端粒酶是一种核糖核蛋白,能以自身RNA模板合成端粒DNA,催化合成TTAGGG重复序列,添加到染色体末端,维持端粒长度不变。端粒酶主要由hTR, TEPl和hTERT组成,一般认为hTERT是端粒酶激活的限速因素。大多数永生化细胞和恶性肿瘤细胞具有端粒酶活性,因而端粒酶目前已成为为细胞持续分裂提供遗传基础的原因。由于端粒酶与细胞衰老、肿瘤发生等关系如此密切,因此已成为肿瘤放射治疗研究热点。目的: 本文利用兰州近代物理研究所重离子研究装置(Heavy Ion Research Facility in Lanzhou,HIRFL)产生的碳离子(31MeV/u 12C6+)对人体细胞、癌细胞进行不同剂量的辐照,探索细胞中端粒酶活性的变化及与之相关的生物学信息。材料与方法: 以人肝细胞HL-7702,肝癌细胞SMMC-7721为实验对象,用不同剂量1Gy、2Gy、3Gy、4Gy的重离子分别对两种细胞进行照射,用多聚酶链式反应-银染端粒重复序列扩增法(PCR- telomeric repeat amplification protocol,TRAP-PCR)检测不同剂量下细胞端粒酶活性的变化。并提取不同剂量下的细胞转入培养皿中,培养10天,固定,染色。统计大于50个细胞的克隆数,绘制细胞存活曲线。结果与讨论: 结果显示,人肝细胞HL-7702自身没有端粒酶活性,经1Gy辐照后也没有端粒酶活性,在2、3Gy处出现端粒酶活性,4Gy处端粒酶活性又消失。肝癌细胞SMMC-7721在1-3Gy处随着剂量的增大端粒酶活性升高,在4Gy处又开始下降;在1-3Gy处随着时间的推移端粒酶活性随着时间而加强(p<0.05)。随着剂量的增大,细胞存活率呈剂量依赖型下降。分析得知,重离子辐射可以诱导人肝细胞产生端粒酶活性,也可以改变肝癌细胞的端粒酶活性。端粒酶参与细胞受辐照后DNA单链损伤的修复;辐照后DNA双链断裂导致端粒酶活性减弱。本实验与其他低LET射线相比,使重离子在辐照治疗中的优势得以体现

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We have developed a new experimental system based on a microfluidic chip to determine severe acute respiratory syndrome coronavirus (SARS-Cov). The system includes a laser-induced fluorescence microfluidic chip analyzer, a glass microchip for both polymerase chain reaction (PCR) and capillary electrophoresis, a chip thermal cycler based on dual Peltier thermoelectric elements, a reverse transcription-polymerase chain reaction (RT-PCR) SARS diagnostic kit, and a DNA electrophoretic sizing kit. The system allows efficient cDNA amplification of SARS-CoV followed by electrophoretic sizing and detection on the same chip. To enhance the reliability of RT-PCR on SARS-CoV detection, duplex PCR was developed on the microchip. The assay was carried out on a home-made microfluidic chip system. The positive and the negative control were cDNA fragments of SARS-CoV and parainfluenza virus, respectively. The test results showed that 17 positive samples were obtained among 18 samples of nasopharyngeal swabs from clinically diagnosed SARS patients. However, 12 positive results from the same 18 samples were obtained by the conventional RT-PCR with agarose gel electrophoresis detection. The SARS virus species can be analyzed with high positive rate and rapidity on the microfluidic chip system.

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The most biological diversity on this planet is probably harbored in soils. Understanding the diversity and function of the microbiological component of soil poses great challenges that are being overcome by the application of molecular biological approaches. This review covers one of many approaches being used: separation of polymerase chain reaction (PCR) amplicons using denaturing gradient gel electrophoresis (DGGE). Extraction of nucleic acids directly from soils allows the examination of a community without the limitation posed by cultivation. Polymerase chain reaction provides a means to increase the numbers of a target for its detection on gels. Using the rRNA genes as a target for PCR provides phylogenetic information on populations comprising communities. Fingerprints produced by this method have allowed spatial and temporal comparisons of soil communities within and between locations or among treatments. Numerous samples can be compared because of the rapid high throughput nature of this method. Scientists now have the means to begin addressing complex ecological questions about the spatial, temporal, and nutritional interactions faced by microbes in the soil environment.

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Temporal trends in total ozone for the St. Lawrence estuary were estimated from ground-based measurements at the NOAA/CMDL station in Caribou, Maine. Linear regression analysis showed that from 1979 to 1999 total ozone has decreased by about 3.3% per decade on an annual basis and ≤6.2% per decade on a monthly basis relative to unperturbed (pre-CFC) levels. The influence of increased ultraviolet-B (280–320 nm) radiation associated with ozone depletion on water column photochemical processes was evaluated by modeling the photobleaching of chromophoric dissolved organic material (CDOM). Linear regression analysis showed small (<0.5% per decade), but statistically significant upward trends in maximum noontime photobleaching rates. Most notably, positive trends in relative rates for May, June, and July, when maximum absolute rates are expected, were predicted. A global model based on TOMS ozone data revealed increases in photobleaching of ≤3% per decade at high latitudes in the Southern Hemisphere. Radiation amplification factors for increases in photochemically weighted UV (280–400 nm) in response to ozone depletion were estimated at 0.1 and 0.08 for photobleaching of CDOM absorbance at 300 and 350 nm, respectively. Application of the laboratory-based model to conditions that more closely resembled those in situ were variable with both overestimation and underestimation of measured rates. The differences between modeled rates and observed rates under quasi-natural conditions were as large or larger than the predicted increases due to ozone depletion. These comparisons suggest that biological activity and mixing play an important, but as yet ill-defined, role in modifying photochemical processes.

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The crystal structure of Er(PM)(3)(TP)(2) [PM = 1-Phenyl-3-methyl-4-isobutyryl-5-pyrazoloiie, TP = triphenyl phosphine oxide] was reported and its photoluminescence properties were studied by UV-vis absorption, excited, and emission spectra. The Judd-ofelt theory was introduced to calculate the radiative transition rate and the radiative decay time of 3.65 ms for the I-4(13/2) -> I-4(15/2) transition of Er3+ ion in this complex.

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Quantitative electrochemilumineseence (ECL) detection of a model protein, bovine serum albumin (BSA) was achieved via biotin-avidin interaction using an avidin-based sensor and a well-developed ECL system of tris(2,2'-bipyridine) ruthenium(II) derivative as label and tri-n-propylamine (TPA) as coreactant. To detect the protein, avidin was linked to the glassy carbon electrode through passive adsorptions and covalent interaction with carboxylate-terminated carbon nanotubes that was used as binder to immobilize avidin onto the electrode. Then, biotinylated BSA tagged with tris(2,2'-bipyridine) ruthenium(II) label was attached to the prepared avidin surface.

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A unique multilabeling at a single-site protocol of the Ru(bpy)(3)(2+) electrochemiluminescence (ECL) system is proposed. Nanoparticles (NPs) were used as assembly substrates to enrich ECL co-reactants of Ru(bpy)(3)(2+) to construct nanoscale-enhanced ECL labels. Two different kinds of NP substrates [including semiconductor NPs (CdTe) and noble metal NPs (gold)] capped with 2-(dimethylamino)ethanethiol (DMAET) [a tertiary amine derivative which is believed to be one of the most efficient of co-reactants of the Ru(bpy)(3)(2+) system] were synthesized through a simple one-pot synthesis method in aqueous media.

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Spherical Ru(bpy)(3)(2+)-doped silica (RuSi) nanoparticles were prepared via a water-in-oil microemulsion approach. The electrochemical and electrochemiluminescent properties of the RuSi nanoparticles immobilized on an indium tin oxide (ITO) electrode were investigated. Further, electrochemiluminescence (ECL) of the RuSi nanoparticles with covalently coated biomacromolecules was studied. By covalent cross-linking with glutaraldehyde, gamma-(aminopropyl) triethoxysilane (APTES)-pretreated RuSi nanoparticles were coupled with different concentrations of bovine serum albumin (BSA), hemoglobin, and myoglobin, respectively.

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An anionic, phosphonate-functionalized polyfluorene, i.e., poly(9,9-bis(3'-phosphatepropyl)fluorene-alt-1,4-phenylene) sodium salt (PFPNa), has been synthesized by copolymerization of phosphonic acid-substituted 2,7-dibromofluorene and phenyldiboronic ester via direct Suzuki polycondensation reaction in DMF/water. Polymer PFPNa is highly soluble and emissive in water with a solubility of 60 mg/mL and a photoluminescence quantum yield of 75%. The absorption and fluorescence spectra of PFPNa are strongly dependent on pH value owing to the partial protonation of phosphate groups and the aggregation of the polymer chains.

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A random lasing emission from 4-(dicy-anomethylene)-2-t-butyl-6(1,1,7,7-tetramethyljulolidyl-9-enyl)-4H-pyran (DCJTB) doped polystyrene (PS) thin films was realized by the scattering role of ZnO nanorods. The device was fabricated by spin-coating DCJTB doped PS on ZnO nanorods. The ZnO nanorods were grown on indium-tin-oxide (ITO) glass substrate by hydrothermal synthesis method. It can be seen that the device emits a resonance multimode peak at center wavelength of 630 nm with a mode line-width of less than 0.23 nm and exhibits threshold excitation intensity as low as 0.375 mJ pulse(-1) cm(-2). The agreement of the dependence of threshold pumped intensity on the excitation area with the random laser theory indicates that the lasing emission realized here is random laser. Our results demonstrate that the nanostructured ZnO nanorods are promising candidate as alternative sources of coherent light emission to realize organic lasers.

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Electrochemistry-based detection methods hold great potential towards development of hand-held nucleic-acid analyses instruments. In this work, we demonstrate the implementation of in situ electrochemical (EC) detection method in a microfluidic flow-through EC-qPCR (FTEC-qPCR) device, where both the amplification of the target nucleic-acid sequence and subsequent EC detection of the PCR amplicon are realized simultaneously at selected PCR cycles in the same device. The FTEC-qPCR device utilizes methylene blue (MB), an electroactive DNA intercalator, for electrochemical signal measurements in the presence of PCR reagent components. Our EC detection method is advantageous, when compared to other existing EC methods for PCR amplicon analysis, since FTEC-qPCR does not require probe-modified electrodes, or asymmetric PCR, or solid-phase PCR. Key technical issues related to surface passivation, electrochemical measurement, PCR inhibition by metal electrode, bubble-free PCR, were investigated. By controlling the concentration of MB and the exposure of PCR mixture to the bare metal electrode, we successfully demonstrated electrochemical measurement of MB in solution-phase, symmetric PCR by amplifying a fragment of lambda phage DNA.

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Herein, we report an approach for protein detection enhanced by ionic liquid (IL) selectors in capillary electrophoresis (CE), with avidin as a model protein. Hydrophilic ILs were added into the running buffer of CE and acted as selectors for sample injection, enriching the positive target and excluding the negative from the capillary. When using 3% (v/v) IL selector, the detection sensitivity of avidin was improved by over one order of magnitude, while the interference from protein adsorption was effectively avoided, even in an uncoated capillary. The electrochemiluminescence method was initially used for IL-based CE with low noise that was independent of the IL concentration, making ILs almost transparent as additives in the electrophoresis buffer.

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Bioactive ultrathin films with the incorporation of amino-terminated G4 PAMAM dendrimers have been prepared via layer-by-layer self-assembly methods on a gold electrode and used for the DNA hybridization analysis. Surface plasmon resonance (SPR), X-ray photoelectron spectroscopy (XPS), and electrochemical impedance spectroscopy (EIS) are used to characterize the successful construction of the multicomponent film on the gold substrate. The dendrimer-modified surfaces improve the immobilization capacity of the probe DNA greatly, compared to the AET (2aminoethanethiol) SAM sensor surfaces without dendrimer molecules. DNA hybridization analysis is monitored by EIS. The dendrimer-based electrochemical impedance DNA biosensor shows high sensitivity and selectivity for DNA hybridization assay. The multicomponent films also display a high stability during repeated regeneration and hybridization cycles.