scholarly journals An Innovative Rapid Thermal Cycling Device for Polymerase Chain Reaction

2008 ◽  
Vol 2 (2) ◽  
Author(s):  
Shadi Mahjoob ◽  
Kambiz Vafai
Keyword(s):  
Heat Transfer ◽  
Polymerase Chain Reaction ◽  
Temperature Distribution ◽  
Thermal Cycling ◽  
Thermal Management ◽  
Uniform Temperature ◽  
Chain Reaction ◽  
Temperature Ramp ◽  
Polymerase Chain ◽  
Thermal Cycler

Polymerase chain reaction (PCR) is the most commonly used molecular biology technique to amplify nucleic acid (DNA and RNA) in vitro. This technique is highly temperature sensitive and thermal management has an important role in PCR operation in reaching the required temperature set points at each step of the process (denaturing, annealing and elongation). In this work, an innovative microfluidic PCR thermal cycling device is designed to increase the heating∕cooling thermal cycling speed while maintaining a uniform temperature distribution throughout the substrate containing the aqueous nucleic acid sample. The device design is incorporating the jet impingement and micro-channel thermal management technologies utilizing a properly arranged configuration filled with a porous medium. Porous Inserts are attractive choices in heat transfer augmentation. They provide a very large surface area for a given volume which is a key parameter in heat transfer processes. Various effective parameters that are relevant in optimizing this flexible thermal cycler are investigated such as thermal cycler configuration, thickness of inlet and exit fluid channels, fluid flow rate and velocity, the porous matrix material and properties, and utilization of thermal grease. An optimized case is established based on the effects of the cited parameters on the temperature ramp, temperature distribution and the required power for circulating the fluid in the thermal cycler. The results indicate that the heating∕cooling temperature ramp (temperature change per heating∕cooling cycling time) of the proposed device is considerably higher (150.82◻C∕s) than those in literature. In addition, the proposed PCR offers a very uniform temperature in the substrate while utilizing a low power.

scholarly journals Design and development of polymerase chain reaction thermal cycler using proportional-integral temperature controller

2018 ◽  
Vol 14 (2) ◽  
pp. 213-218
Author(s):  
Chong Kim Soon ◽  
Nawoor Anusha Devi ◽  
Kok Beng Gan ◽  
Sue-Mian Then
Keyword(s):  
Polymerase Chain Reaction ◽  
Low Cost ◽  
Maximum Temperature ◽  
Gradient System ◽  
Chain Reaction ◽  
Temperature Controller ◽  
Proportional Integral ◽  
Integral Temperature ◽  
Polymerase Chain ◽  
Thermal Cycler

A thermal cycler is used to amplify segments of DNA using the polymerase chain reaction (PCR). It is an instrument that requires precise temperature control and rapid temperature changes for certain experimental protocols. However, the commercial thermal cyclers are still bulky, expensive and limited for laboratory use only.  As such it is difficult for on-site molecular screening and diagnostics. In this work, a portable and low cost thermal cycler was designed and developed. The thermal cycler block was designed to fit six microcentrifuge tubes. A Proportional-Integral temperature controller was used to control the thermal cycler block temperature. The results showed that the maximum temperature ramp rate of the developed thermal cycler was 5.5 °C/s. The proportional gain (Kp) and integral gain (Ki) of the PI controller were 15 A/V and 1.8 A/Vs respectively. Finally, the developed thermal cycler successfully amplified six DNA samples at the expected molecular weight of 150 base pair. It has been validated using the Eppendorf Mastercycler nexus gradient system and gel electrophoresis analysis

scholarly journals Intracellular amplification of proviral DNA in tissue sections using the polymerase chain reaction.

1992 ◽  
Vol 40 (3) ◽  
pp. 333-341 ◽  
Author(s):  
K P Chiu ◽  
S H Cohen ◽  
D W Morris ◽  
G W Jordan
Keyword(s):  
Polymerase Chain Reaction ◽  
Cultured Cells ◽  
Chain Reaction ◽  
Tissue Sections ◽  
Mouse Mammary Tumor ◽  
Tumor Virus ◽  
Polymerase Chain ◽  
Thermal Cycler ◽  
Cellular Dna

We developed a new method to amplify cell DNA in situ using the polymerase chain reaction (PCR). Proviral sequences of mouse mammary tumor virus (MMTV) contained in cultured cells and tissue sections were amplified intracellularly using a thermal cycler. Two techniques were employed to maintain the localization of the amplified DNA. First, complementary tails at the 5' ends of the oligonucleotide primers resulted in the synthesis of high molecular weight concatamers containing the target sequences. Second, the PCR was carried out in a thin film of agarose solidified over the tissue sections. The specifically amplified and localized DNA was then detected by in situ hybridization (ISH). Our results demonstrate that (a) DNA in tissue sections can serve as the target for the polymerase chain reaction in situ, (b) cell morphology is maintained, and (c) a target of 167 BP can be specifically detected in individual cells. This technique should be generally applicable to amplifying cellular DNA targets in tissue sections for detection in situ.

A Thermal System for a High Throughput Continuous Flow Polymerase Chain Reaction Device (CFPCR)

2008 ◽  
Author(s):  
P.-C. Chen ◽  
D. S. Park ◽  
B. H. You ◽  
N. Kim ◽  
T. Park ◽  
...  
Keyword(s):  
Temperature Distribution ◽  
High Throughput ◽  
Continuous Flow ◽  
Thermal Environment ◽  
Infrared Camera ◽  
Thermal System ◽  
Uniform Temperature ◽  
Chain Reaction ◽  
Copper Strip ◽  
Polymerase Chain

A thermal system used to evaluate a high throughput 96 continuous flow polymerase chain reactor (CFPCR) array was designed, fabricated, and tested. Each polymerase chain reactor (PCR) in the array required three different temperature zones to realize denaturaiton at 90°C–94°C, renaturation at 50°C–70°C, and extension at 72°C; a total of 288 temperature zones were required for the 96 CFPCR array. In an initial configuration, 18 copper strips were used to define the 288 temperature zones. Each copper strip was controlled by a PID feedback control loop. Numerical simulations were used to understand the thermal crosstalk phenomena between the micromilled copper strips, which were tightly packed since the high throughput micro-titer plate format restricted each CFPCR to a square 8 mm on a side. The lowest achievable temperature in each renaturation zone in this complicated thermal environment was also identified. Thermal crosstalk limited the minimum renaturation temperature to 61.1°C. An infrared camera was used to investigate the temperature uniformity over a 0.25 mm thick polycarbonate sheet mounted on the thermal system. The temperature distribution was not uniform due to poor contact between the copper strips and device, warm air accumulated between the packed copper strips, and greater heat transfer around the boundaries of the device. More work is required to overcome these limitations and achieve a more uniform temperature distribution for a multi well CFPCR.

scholarly journals Specific Detection of Clostridium botulinum Types A, B, E, and F Using the Polymerase Chain Reaction

2002 ◽  
Vol 85 (5) ◽  
pp. 1025-1028 ◽  
Author(s):  
Kathy E Craven ◽  
Joseph L Ferreira ◽  
Mark A Harrison ◽  
Paul Edmonds
Keyword(s):  
Polymerase Chain Reaction ◽  
Clostridium Botulinum ◽  
Extraction Procedure ◽  
Toxin Gene ◽  
Chain Reaction ◽  
Gene Target ◽  
Human Illness ◽  
Polymerase Chain ◽  
Clostridial Species ◽  
Thermal Cycler

Abstract Clostridium botulinum organisms generally produce 1 of 4 neurotoxin types (A, B, E, and F) associated with human illness. Neurotoxin type determination is important in identification of the bacterium. A polymerase chain reaction (PCR) method was developed to identify 24 h botulinal cultures as potential types A, B, E, and F neurotoxin producers as well as other clostridial species which also produce neurotoxins. Components of the PCR and amplification conditions were adjusted for optimal amplification of toxin gene target regions to enable simultaneous testing for types A, B, E, and F in separate tubes using a single thermal cycler. Each primer set was specific for its corresponding toxin type. A DNA extraction procedure was also included to remove inhibitory substances that may affect amplification. This procedure is rapid, sensitive, and specific for identification of toxigenic C. botulinum.

Effect of convection modes on the polymerase chain reaction in a square cavity

2012 ◽  
Vol 9 (2) ◽  
pp. 86-89
Author(s):  
K.V. Moiseyev
Keyword(s):  
Heat Transfer ◽  
Polymerase Chain Reaction ◽  
Concentration Field ◽  
Boussinesq Approximation ◽  
Convection Flow ◽  
Square Cavity ◽  
Chain Reaction ◽  
Physicochemical Interactions ◽  
Polymerase Chain ◽  
Substance Transfer

The effect of various regimes of free-convective heat transfer of a Newtonian incompressible fluid on the polymerase chain reaction in a square cavity is numerically studied. As a mathematical model of the process of free convection, the Oberbeck-Boussinesq approximation is considered. For the polymerase chain reaction, the equations of substance transfer are used, taking into account the physicochemical interactions of the components of the mixture. It was believed that the mixture consists of three components. The results of calculations allow us to estimate the effect of convection flow regimes on the overall duration of the reaction and the concentration field of the components, and also to determine the optimum thermal parameters for PCR.

Failure Prediction of Polymerase Chain Reaction Thermal Cycler

2017 ◽  
Author(s):  
Mi-So Lee ◽  
Chan-Young Park ◽  
Yu-Seop Kim ◽  
Hye-Jeong Song ◽  
Jong-Dae Kim
Keyword(s):  
Polymerase Chain Reaction ◽  
Failure Prediction ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Thermal Cycler
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