Continuous-flow Polymerase Chain Reaction Chip Research Articles

A Portable Continuous-Flow Polymerase Chain Reaction Chip Device Integrated with Arduino Boards for Detecting Colla corii asini.

Food security is a significant issue in modern society. Because morphological characters are not reliable enough to distinguish authentic traditional Chinese medicines, it is essential to establish an effective and applicable method to identify them to protect people’s health. Due to the expensive cost of the manufacturing process and the large volume of the analytical system, the need to build a portable and cheap device is urgent. This work describes the development of a portable nucleic acid amplification device integrated with thermal control and liquid pumping connecting to Arduino boards. We present a novel microfluidic polymerase chain reaction (PCR) chip with symmetric isothermal zones. The total chip volume is small, and only one Arduino board is needed for thermal control. We assemble a miniaturized liquid pump and program an Arduino file to push the sample mixture into the chip to implement the PCR process. In the proposed operation, the Nusselt number of the sample flow is less than one, and the heat transfer is conduction only. Then we can ensure temperature uniformity in specific reaction regions. A Colla corii asini DNA segment of 200 bp is amplified to evaluate the PCR performance under the various operational parameters. The initial concentration for accomplishing the PCR process is at least 20 ng/μL at the flow rate of 0.4 μL/min in the portable continuous flow PCR (CFPCR) device. To our knowledge, our group is the first to introduce Arduino boards into the heat control and sample pumping modules for a CFPCR device.

Advance in Research of Microfluidic Polymerase Chain Reaction Chip

Abstract Gene is the genetic code of human beings, and only 0.1‰ difference among individuals but creates a wealthy diversity. Thus it is significant in practicality to explore the difference. Polymerase chain reaction (PCR) is one of the technologies mostly used in genetic research. But traditional thermal cyclers are not only time- and energy-consuming but also difficult to be integrated and portable. The combination of microfluidics and PCR can lessen the reaction volume significantly and increase the reaction efficiency with easy integration and miniaturization. In this review, we briefly introduced microchamber PCR chip and continuous-flow PCR chip according to their structures, as well the detection methods including capillary electrophoresis, fluorescence, electrochemistry and DNA hybridization array. Finally, we summarized the recent advance and pointed out the future development of microfluidic PCR chip.

All-plastic, low-power, disposable, continuous-flow PCR chip with integrated microheaters for rapid DNA amplification

The design, fabrication and evaluation of a low-cost and low-power, continuous-flow microfluidic device for DNA amplification by polymerase chain reaction (PCR) with integrated heating elements, on a commercially available thin polymeric substrate (Pyralux® Polyimide), is presented. The small thermal mass of the chip, in combination with the low thermal diffusivity of the polymeric substrate on which the heating elements reside, yields a low power consumption PCR chip with fast amplification rates. A flow-through μPCR device is designed and fabricated using flexible printed circuit (FPC) technology on a foot-print area of 8cm×6cm with a meandering microchannel realized at a very small distance (50μm) above 3 independently operating resistive (copper) serpentine microheaters, each one defining one of the three PCR temperature zones. The 145cm-long microchannel is appropriately designed to cross the alternating temperature zones as many times as necessary for the DNA sample to perform 30 PCR cycles. Numerical computations lead the design so that there is no thermal crosstalk between the 3 zones of our chip and indicate excellent temperature uniformity in each zone. In addition, the total power consumption during the chip operation is calculated to be in the order of a few Watts, verified experimentally by means of thermal characterization of our heaters. Thermal camera measurements also verified the excellent temperature uniformity in the three thermal zones. An external, home-made temperature control system was utilized to maintain the heater temperatures in the designated values (±0.2°C). The PCR chip was validated by a successful amplification of a 90 base-pairs DNA template of the mouse GAPDH housekeeping gene within 5min.

MCU based real-time temperature control system for universal microfluidic PCR chip

The analysis of genetic materials in the post-human genome project era has become an ever-expanding branch of research and thus routinely employed in majority of biochemical laboratories. Most of the diagnostic research area emphasizes on polymerase chain reaction for detecting pathogenic organisms. However, the conventional polymerase chain reaction requires expensive and sophisticated thermal cycler and is not handy owing to its large dimensions. Therefore, we fabricated a continuous-flow polymerase chain reaction chip on a PDMS based microfluidic platform to ease the hardship of the conventional system. Temperature being the most crucial factor in polymerase chain reaction, was monitored and regulated by thermostatic action using an on-line computer system. Indium tin oxide coated glass platform was used for heating as it is transparent and has good thermal conductivity under the influence of DC bias. The heating circuit used an ATMega 128 MCU to control the temperature. As a result, a precise and quick heating environment was maintained on the microfluidic chip to amplify the target DNA. We successfully amplified Lambda phage and Escherichia coli DNA on our chip to prove the practicality of the device.

Computer Simulation Analysis of Microchannel for a Continuous-Flow PCR Chip

The purpose of this study is to analyze the model of a continuous-flow Polymerase Chain Reaction (PCR) chip with CoventorWare software. It offered the ability of fast heating and cooling rate when the DNA duplication process happened. The main idea was to compare the effects of different cross sections of channel and find out the better one for heat transfer.

Continuous-Flow Polymerase Chain Reaction Chip by Water Cooling

A novel continuous-flow polymerase chain reaction (PCR) chip has been analyzed. It operates by cycling a prepared sample within three temperature zones. Two temperature zones are controlled by two thermal controllers and the other is controlled by the flow rate of the fluid inside a cooling channel under a glass chip. The chip system consists of the reaction PDMS chip, a cover glass chip, a cooling channel, and is equipped with cartridge heaters and thermocouples for temperature control. Commercial software is utilized to determine the chip materials that are responsible for creating the denaturation, annealing and extension temperature zones within the chip. Therefore we utilize this chip to perform PCR experiments. Results show that DNA templates are amplified successfully.

Continuous-Flow Polymerase Chain Reaction Chip by Water Cooling

Continuous-Flow Polymerase Chain Reaction Chip by Water Cooling

Glass-Based Continuous-Flow PCR Chip With a Portable Control System for DNA Amplification

A glass-based continuous-flow polymerase chain reaction (PCR) chip has been designed and fabricated. The device consists of a glass microfluidic channel, three NiCr heaters, and three Ni thermometers on the silicon substrate. An intelligent temperature-control circuit system has been designed to achieve desirable temperature control (95, 72, and 55 <formula formulatype="inline" xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink"><tex Notation="TeX">$^{\circ}{\rm C}$</tex></formula> ) at the three temperature zones of the PCR chip. Simulation underneath the microfluidic channel using the finite element method shows that the temperature distribution through the three temperature zones are relatively uniform. A mixture of DNA samples for PCR was allowed to flow through the microfluidic channel under different flow rates. The amplified sample of the target DNA obtained from the PCR chip was then separated by electrophoresis and was analyzed using an ultraviolet analyzer. The result indicates that DNA amplification can be achieved and that its amplification factor depends greatly on the injection rate of the sample. The optimum sample-flow rate is 0.6 <formula formulatype="inline" xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink"><tex Notation="TeX">$\mu{\rm l}/{\rm min}$</tex></formula> .

Kinetic characteristics of continuous flow polymerase chain reaction chip: A numerical investigation

Continuous flow PCR (polymerase chain reaction) chip holds impressive advantages compared to micro chamber PCR chip. In order to have better understanding of kinetic characteristics of continuous flow PCR chip, a comprehensive mathematical model is presented in this paper, including melting, annealing and extension phases of a typical PCR process which has the essence of a convection-diffusion-reaction system. Using this model, we can simulate the PCR process in series of reaction cycles. Numerical results show that the average sample velocity plays a significant role in affecting the amplification efficiency. Also, appropriate combination of the PCR mixture is important for high-quality DNA amplification. Giving a large initial DNA concentration range, the continuous flow PCR scheme holds excellent real-time detection ability theoretically. The present numerical model bridges the temperature distribution to the real DNA amplification, and thereby is able to successfully predict continuous flow PCR properties which are important for the chip design.

Series DNA Amplification Using the Continuous-Flow Polymerase Chain Reaction Chip

We proposed a continuous-flow polymerase chain reaction (PCR) chip that can be used for series DNA amplification. The continuous-flow PCR chip has several advantages such as fast thermal cycling, series of amplifications, cost-effective fabrication, portability, and fluorescence detection. The continuous-flow PCR chip is composed of two parts namely poly(dimethylsiloxane) (PDMS) microchannel for sample injection and indium–tin-oxide (ITO) heater/glass chip for thermal cycling. The fabricated microchannel width and depth are 250 and 200 µm, respectively. Also, the total working length of the PDMS microchannel is 1340 mm which is equivalent for 20 cycles of amplification. A 2:2:3 microchannel length ratio for three different temperature zones namely denaturation, annealing, and extension was assigned, respectively. Upon the operation of the fabricated continuous-flow PCR chip, the amplification of plasmid DNA pKS-GFP with 720 base pairs and PG-noswsi with 300 base pairs were found successfully with a total reaction time of 15 min.

Fabrication and characterization of a PDMS–glass hybrid continuous-flow PCR chip

A continuous-flow (CF) microchip was fabricated using polydimethylsiloxane (PDMS)/glass laminate for efficient polymerase chain reaction (PCR). The temperatures of the three separate zones, corresponding to the three PCR thermal cycling steps were controlled using an external PID temperature controller. A PCR mixture was introduced to the CF microchip using a syringe pump. The hydrophobic surface of the PDMS channel which adsorbs enzymes was modified by adding 2.5% polyvinylpyrrolidone (PVP) to the PCR mixture. The effect of flow rate in the CF chip on DNA amplification was examined and its PCR efficiency was compared with that of conventional PCR. The microchip requires only 8–30 min to produce detectable amounts of PCR products as determined by slab gel separation, whereas a conventional thermocycler requires 90–150 min over 30 amplification cycles to achieve the same result.