scholarly journals Snapback Primer Genotyping with Saturating DNA Dye and Melting Analysis

2008 ◽  
Vol 54 (10) ◽  
pp. 1648-1656 ◽  
Author(s):  
Luming Zhou ◽  
Roscoe J Errigo ◽  
Hongzhe Lu ◽  
Mark A Poritz ◽  
Michael T Seipp ◽  
...  
Keyword(s):  
Full Length ◽  
Stem Length ◽  
Clinical Samples ◽  
Physical Separation ◽  
Dna Hairpins ◽  
Pcr Products ◽  
Melting Analysis ◽  
Higher Temperature ◽  
Covalent Modifications ◽  
Lower Temperature

Abstract Background: DNA hairpins have been used in molecular analysis of PCR products as self-probing amplicons. Either physical separation or fluorescent oligonucleotides with covalent modifications were previously necessary. Methods: We performed asymmetric PCR for 40–45 cycles in the presence of the saturating DNA dye, LCGreen Plus, with 1 primer including a 5′ tail complementary to its extension product, but without any special covalent modifications. Samples were amplified either on a carousel LightCycler for speed or on a 96/384 block cycler for throughput. In addition to full-length amplicon duplexes, single-stranded hairpins were formed by the primer tail “snapping back” and hybridizing to its extension product. High-resolution melting was performed on a HR-1 (for capillaries) or a LightScanner (for plates). Results: PCR products amplified with a snapback primer showed both hairpin melting at lower temperature and full-length amplicon melting at higher temperature. The hairpin melting temperature was linearly related to the stem length (6–28 bp) and inversely related to the log of the loop size (17–135 bases). We easily genotyped heterozygous and homozygous variants within the stem, and 100 blinded clinical samples previously typed for F5 1691G>A (Leiden) were completely concordant by snapback genotyping. We distinguished 7 genotypes in 2 regions of CFTR exon 10 with symmetric PCR using 2 snapback primers followed by product dilution to favor intramolecular hybridization. Conclusions: Snapback primer genotyping with saturating dyes provides the specificity of a probe with only 2 primers that are free of special covalent labels in a closed-tube system.

scholarly journals Sprouting of Sorghum (Sorghum bicolor [L.] Moench): Effect of Drying Treatment on Protein and Starch Features

Foods ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 407 ◽  
Author(s):  
Mia Marchini ◽  
Alessandra Marti ◽  
Claudia Folli ◽  
Barbara Prandi ◽  
Tommaso Ganino ◽  
...  
Keyword(s):  
Food Production ◽  
Cost Effective ◽  
High Exposure ◽  
Effective Technology ◽  
Drying Treatment ◽  
Higher Temperature ◽  
Lower Temperature ◽  
The One ◽  
Drying Conditions ◽  
Sorghum Bicolor L

The nutritional and physicochemical properties of sorghum proteins and starch make the use of this cereal for food production challenging. Sprouting is a cost-effective technology to improve the nutritional and functional profile of grains. Two drying treatments were used after sorghum sprouting to investigate whether the drying phase could improve the protein and starch functionalities. Results showed that the drying treatment at lower temperature/longer time (40 °C for 12 h) extended the enzymatic activity that started during sprouting compared to the one performed at higher temperature/shorter time (50 °C for 6 h). An increased protein hydrolysis and water- and oil-holding capacity were found in the flour obtained by the former treatment. Higher protein matrix hydrolysis caused high exposure of starch to enzymes, thus increasing its digestibility, while worsening the technological functionality. Overall, modulating drying conditions could represent a further way, in addition to sprouting, to improve sorghum flour’s nutritional profile.

MICROSTRUCTURAL EVOLUTION OF Al-Cu-Mg-Ag ALLOY WITH TRACE Zr ADDITION DURING TWO-STEP HOMOGENIZATION

2009 ◽  
Vol 23 (06n07) ◽  
pp. 855-862 ◽  
Author(s):  
FEIYUE MA ◽  
ZHIYI LIU
Keyword(s):  
Melting Point ◽  
Grain Boundaries ◽  
Microstructural Evolution ◽  
Cast Alloy ◽  
Scanning Calorimetry ◽  
Zr Addition ◽  
Homogenization Time ◽  
The Matrix ◽  
Higher Temperature ◽  
Lower Temperature

The microstructural evolution in an Al - Cu - Mg - Ag alloy with trace Zr addition during homogenization treatment was characterized by Differential Scanning Calorimetry (DSC), Scanning Electron Microscopy (SEM) and Energy-dispersive X-ray Spectroscopy (EDS). It was shown that the low-melting-point phase segregating toward grain boundaries is Al 2 Cu , with a melting point of 523.52°C. A two-step homogenization process was employed to optimize the microstructure of the as-cast alloy, during which the alloy was first homogenized at a lower temperature, then at a higher temperature. After homogenized at 420°C for 6 h, Al 3 Zr particles were finely formed in the matrix. After that, when the alloy was homogenized at an elevated temperature for a longer time, i.e., 515°C for 24 h, most of the precipates at the grain boundaries were removed. Furthermore, the dispersive Al 3 Zr precipitates were retained, without coarsening greatly in the final homogenization step. A kinetics model is employed to predict the optimal homogenization time at a given temperature theoretically, and it confirms the result in present study, which is 420°C/6h+515°C/24h.

scholarly journals Research on the Static Recrystallization and Precipitation Behaviors of a V-N Microalloyed Steel

2013 ◽  
Vol 2013 ◽  
pp. 1-7 ◽  
Author(s):  
Baochun Zhao ◽  
Tan Zhao ◽  
Guiyan Li ◽  
Qiang Lu
Keyword(s):  
Static Recrystallization ◽  
Microalloyed Steel ◽  
Time Interval ◽  
Main Role ◽  
Recrystallization Process ◽  
Compression Tests ◽  
Double Compression ◽  
Higher Temperature ◽  
Lower Temperature ◽  
Temperature Strain

Double compression tests were performed on a Gleeble-3800 thermomechanical simulator to study the softening behaviors of deformed austenite in a V-N microalloyed steel. The static recrystallization volume fractions were calculated by stress offset method, and the kinetic model of static recrystallization was constructed. The effects of temperature, strain, and time interval on the softening behaviors were analyzed, and the interactions between precipitation and recrystallization were discussed. The results show that the softening behaviors of the deformed austenite at lower temperature or higher temperature are markedly different. At the temperature of 850°C or 800°C, pinning effects of the precipitates play the main role, and the recrystallization process is inhibited, which leads to the formation of plateaus in the softening curves. An increase in strain promotes the precipitation and recrystallization processes while reduces the inhibition effect of precipitation on recrystallization as well.

Effects of Sr Addition on Mechanical Properties and Microstructure of Mg-6Al Magnesium Alloy

2011 ◽  
Vol 686 ◽  
pp. 120-124
Author(s):  
Jin Ping Fan ◽  
She Bin Wang ◽  
Bing She Xu
Keyword(s):  
Mechanical Properties ◽  
Tensile Strength ◽  
Magnesium Alloy ◽  
Strengthening Mechanism ◽  
Microstructure And Mechanical Properties ◽  
Higher Temperature ◽  
Lower Temperature

The effects of Sr addition on the mechanical properties and microstructure of Mg-6Al mag- nesium alloy both at 25 °C and at 175 °C were investigated by means of OM, SEM and EDS and XRD. Upon the Sr addition of 2%, the tensile strength was increased by 7.2% to 184.4MPa at 25 °C, while it was increased by 30% to 155.4MPa at 175 °C. The strengthening mechanism of Mg-6Al-xSr at lower temperature (25 °C) was different from that at higher temperature (175°C). The results show that the addition of strontium effectively improved the microstructure and mechanical properties of magnesium alloy.

The Effect of Uniaxial Cyclic Deformation on the Evolution of Phase Transformation Fronts in Pseudoelastic NiTi Wire

2001 ◽  
Author(s):  
Mark A. Iadicola ◽  
John A. Shaw
Keyword(s):  
Mechanical Response ◽  
Infrared Imaging ◽  
Partial Transformation ◽  
Niti Wire ◽  
Full Field ◽  
Temperature Experiment ◽  
Background Temperature ◽  
Higher Temperature ◽  
Lower Temperature ◽  
Niti Wires

Abstract Experiments are presented of the response of pseudoelastic NiTi wires subjected to displacement controlled cycles. A custom built thermo-mechanical testing apparatus is used to control the background temperature field of the wire specimen while allowing the evolution of transformation fronts to be tracked by full field infrared imaging. Two experiments under similar end-displacement histories, but at temperatures ≈8°C apart, are shown to give remarkably different cyclic responses. The mechanical response for the lower temperature experiment continued to soften but retained its shape through 43 partial transformation cycles, and the pattern of transformation fronts seemed to reach a steady state. The response for the higher temperature experiment showed a change in shape of the mechanical response and distinct changes in transformation front patterns over 31 partial transformation cycles.

A Numerical Analysis of Gas Turbine Disks Incorporating Rotating Heat Pipes

2000 ◽  
Author(s):  
Y. Cao ◽  
J. Ling ◽  
R. Rivir ◽  
C. MacArthur
Keyword(s):  
Heat Pipe ◽  
Gas Turbine ◽  
Heat Pipes ◽  
High Heat ◽  
Thermal Dissipation ◽  
High Heat Flux ◽  
Heating And Cooling ◽  
Turbine Disks ◽  
Higher Temperature ◽  
Lower Temperature

Abstract Radially rotating heat pipes have been proposed for cooling gas turbine disks working at high temperatures. A disk incorporating the heat pipe would have an enhanced thermal dissipation capacity and a much lower temperature at the disk rim and dovetail surface. In this paper, extensive numerical simulations have been made for heat-pipe-cooled disks. Thermal performances are compared for the disks with and without incorporating the heat pipe at different heating and cooling conditions. The numerical results presented in this paper indicate that radially rotating heat pipes can significantly reduce the maximum and average temperatures at the disk rim and dovetail surface under a high heat flux working condition. In general, the maximum and average temperatures at the disk rim and dovetail surface could be reduced by above 250 and 150 degrees, respectively, compared to those of the disk without the heat pipe. As a result, a disk incorporating radially rotating heat pipes could alleviate temperature-related problems and allow a gas turbine to work at a much higher temperature.

scholarly journals Integrated Extreme Real-Time PCR and High-Speed Melting Analysis in 52 to 87 Seconds

2019 ◽  
Vol 65 (2) ◽  
pp. 263-271 ◽  
Author(s):  
Joseph T Myrick ◽  
Robert J Pryor ◽  
Robert A Palais ◽  
Sean J Ison ◽  
Lindsay Sanford ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
High Speed ◽  
Dna Analysis ◽  
Point Of Care ◽  
Turnaround Time ◽  
Single Nucleotide Variants ◽  
Cycle Times ◽  
Pcr Products ◽  
Melting Analysis

Abstract BACKGROUND Extreme PCR in <30 s and high-speed melting of PCR products in <5 s are recent advances in the turnaround time of DNA analysis. Previously, these steps had been performed on different specialized instruments. Integration of both extreme PCR and high-speed melting with real-time fluorescence monitoring for detection and genotyping is presented here. METHODS A microfluidic platform was enhanced for speed using cycle times as fast as 1.05 s between 66.4 °C and 93.7 °C, with end point melting rates of 8 °C/s. Primer and polymerase concentrations were increased to allow short cycle times. Synthetic sequences were used to amplify fragments of hepatitis B virus (70 bp) and Clostridium difficile (83 bp) by real-time PCR and high-speed melting on the same instrument. A blinded genotyping study of 30 human genomic samples at F2 c.*97, F5 c.1601, MTHFR c.665, and MTHFR c.1286 was also performed. RESULTS Standard rapid-cycle PCR chemistry did not produce any product when total cycling times were reduced to <1 min. However, efficient amplification was possible with increased primer (5 μmol/L) and polymerase (0.45 U/μL) concentrations. Infectious targets were amplified and identified in 52 to 71 s. Real-time PCR and genotyping of single-nucleotide variants from human DNA was achieved in 75 to 87 s and was 100% concordant to known genotypes. CONCLUSIONS Extreme PCR with high-speed melting can be performed in about 1 min. The integration of extreme PCR and high-speed melting shows that future molecular assays at the point of care for identification, quantification, and variant typing are feasible.

The Self-Decay Characteristics of Chlorine Dioxide in Water

2012 ◽  
Vol 610-613 ◽  
pp. 296-299
Author(s):  
Xin Jie Li ◽  
Dan Nan Jiang ◽  
Yue Jun Zhang
Keyword(s):  
Rate Constants ◽  
Tap Water ◽  
Pure Water ◽  
Decay Behaviour ◽  
Increase With Increase ◽  
Higher Temperature ◽  
Effects Of Temperature ◽  
Lower Temperature ◽  
Kinetics Of ◽  
Temperature Water

In order to learn the ClO2 decay behaviour in tap water, the kinetics of ClO2 decay in pure water was studied. Under the conditions of tap water treatment and keeping away from light, the effects of temperature and pH on ClO2 degradation were investigated. The results show that the ClO2 decay reaction in pure water is the first-order with respect to ClO2, the decay rate constants increase with increase in temperature or pH. At pH=6.87, the rate constants are 0.012h-1(15°C), 0.017h-1(25°C), 0.023h-1(35°C), and 0.029h-1(45°C), respectively. At 25°C, the rate constants are 0.0083h-1(pH=4.5), 0.0111h-1(pH=5.5), 0.0143h-1(pH=6.5), 0.0222h-1(pH=7.5), and 0.0351h-1(pH=8.5), respectively. The experimental data prove that ClO2 is more stable in acidic or lower temperature water than in neutral, alkalescent, or higher temperature water.

scholarly journals Simplified Fluorescent Multiplex PCR Method for Evaluation of the T-Cell Receptor Vβ-Chain Repertoire

2005 ◽  
Vol 12 (4) ◽  
pp. 477-483 ◽  
Author(s):  
Sanjit Fernandes ◽  
Surendra Chavan ◽  
Vivek Chitnis ◽  
Nina Kohn ◽  
Savita Pahwa
Keyword(s):  
T Cell ◽  
Multiplex Pcr ◽  
T Cell Receptor ◽  
Cell Receptor ◽  
T Cell Subsets ◽  
Clinical Samples ◽  
Cdr3 Length ◽  
Pcr Products ◽  
Pcr Method ◽  
T Cell Receptor Vβ

ABSTRACTRationale: evaluation of the T-cell receptor (TCR) Vβ-chain repertoire by PCR-based CDR3 length analysis allows fine resolution of the usage of the TCR Vβ repertoire and is a sensitive tool to monitor changes in the T-cell compartment. A multiplex PCR method employing 24 labeled upstream Vβ primers instead of the conventionally labeled downstream Cβ primer is described. Method: RNA was isolated from purified CD4 and CD8 T-cell subsets from umbilical cord blood and clinical samples using TRI reagent followed by reverse transcription using a Cβ primer and an Omniscript RT kit. The 24 Vβ primers were multiplexed based on compatibility and product sizes into seven reactions. cDNA was amplified using 24 Vβ primers (labeled with tetrachloro-6-cardoxyfluorescein, 6-carboxyfluorescein, and hexachloro-6-carboxyfluorescein), an unlabeled Cβ primer, and Taqgold polymerase. The fluorescent PCR products were resolved on an automated DNA sequencer and analyzed using the Genotyper 2.1 software. Results: Vβ spectratypes of excellent resolution were obtained with RNA amounts of 250 ng using the labeled Vβ primers. The resolution was superior to that obtained with the labeled Cβ primer assay. Also the numbers of PCRs were reduced to 7 from the 12 required in the Cβ labeling method, and the sample processing time was reduced by half. Conclusion: The method described for T-cell receptor Vβ-chain repertoire analysis eliminates tedious dilutions and results in superior resolution with small amounts of RNA. The fast throughput makes this method suitable for automation and offers the feasibility to perform TCR Vβ repertoire analyses in clinical trials.

scholarly journals The Investigation of Viscometric Properties of the Most Reputable Types of Viscosity Index Improvers in Different Lubricant Base Oils: API Groups I, II, and III

Lubricants ◽  
2022 ◽  
Vol 10 (1) ◽  
pp. 6
Author(s):  
Seyed Ali Khalafvandi ◽  
Muhammad Ali Pazokian ◽  
Ehsan Fathollahi
Keyword(s):  
Viscosity Index ◽  
Behavioral Differences ◽  
Base Oil ◽  
Polymer Molecules ◽  
Base Oils ◽  
Two Temperatures ◽  
Higher Temperature ◽  
Viscosity Index Improvers ◽  
Lower Temperature ◽  
Definition Of

Four commercial viscosity index improvers (VII) have been used to investigate the behavioral differences of these compounds in three types of universally applicable base oils. The used VIIs are structurally three types of co-polymer: ethylene-propylene, star isoprene, and two di-block styrene-isoprene. After dissolving of different amounts of VIIs in different base oils, the kinematic viscosities at two standard temperatures were determined and the intrinsic viscosities were calculated according to Huggins method, then the effects of changes in base oil and polymer type were investigated. Intrinsic viscosities as criteria for polymer molecules sizes were found to be higher at lower temperature than at higher temperature. Dependence of intrinsic viscosity on the polymer molecular weight was observed. In the previous works, one or two types of VIIs were studied in only one type of base oil and/or solvent, not different base oils. Furthermore, different ranges of temperatures and concentrations not necessarily applied ranges were selected, but in this work, common base oils and most commercial VIIs were used and the viscometric properties were compared at two temperatures. Viscosities at these temperatures are used for determining VI and definition of lubricant’s viscosity grades. VI improvement is the main cause of VII usage.

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