Miniaturized and IoT enabled Continuous-flow based Microfluidic PCR Device for DNA Amplification

Abstract : Herein, a continuous-flow driven microfluidic device has been designed and fabricated using the CO2 laser ablation method for polymerase chain reaction (PCR). The device consists of a polymethyl methacrylate (PMMA) microfluidic channel with 30 serpentine thermal cycles, an arduino board, two custom-made cartridge heaters, and thermocouple sensors. The portable thermal management system, with aluminium blocks placed on a wooden substrate, working on the PID controller principle, is low-cost, battery-powered, automated, integrated, and IoT-enabled. The device with dimensions 80 × 72 × 36 mm3 (L x W x H) has a temperature accuracy of ±0.2°C. The IoT module enables accessing and storage of real-time temperature values directly onto the smartphone through ThingSpeak analytics. It was developed to achieve desirable accurate temperature at two thermal zones, denaturation and annealing (95°C and 60°C) on the microfluidic thermal management platform. A PCR mixture of 20 µ l was infused into the serpentine-based microchannel using a syringe pump. Amplification of DNA template with 594-base pair (bp) fragment of the rat GAPDH gene was successfully performed on the miniaturized thermal management system. The total time required for a complete PCR reaction was 32 min at an optimum flow rate of 5 µ l/min. The amplified sample of the target DNA obtained from the PCR microchannel was then separated by agarose gel electrophoresis and was further analyzed using a gel-doc system. Finally, the obtained results were compared to the conventional PCR instrument showing excellent performance.
 EXISTING SYSTEM :
 ? The particular optimized heater spacings were derived by instilling a zero heat flux boundary condition in the y-direction at the leading edges of each of the heaters. Due to the 96 C heater, a negative temperature gradient exists in the positive x direction. ? If a reasonable pressure drop exists in the downstream direction, then it is highly improbable that flow recirculation will occur. ? Oppositely, adverse pressure gradients (backpressure) are much more likely to induce some recirculation regimes near the microchannel wall. ? It was found in the literature that when the angles of the converging/diverging conduits were small, the probability of flow recirculation is minimized vis-à-vis steep profiles.
 DISADVANTAGE :
 ? In order to solve the above problems, a novel digital PCR system was proposed in the present work. ? The problem of detecting data failures can also serve as a reagent for screening dimers in the assay. ? To avoid PCR inhibition, it is important to prevent the direct interaction between PCR mixtures and device surfaces. However, the use of plugs/segments does not eliminate the adsorption problem even with a surface treatment. ? Therefore early droplet-based microfluidic PCR devices emerged to tackle the contamination problem. ? Future smart microfluidic platform will have a major impact on artificial intelligence and biological computations.
 PROPOSED SYSTEM :
 • The proposed device was applicable both in the material science laboratory and in the synthesis industry. • The proposed device needs other equipment such asmicro-pump stability control makes the process, thus it is costlier and difficult to operatefor minuscule synthesis. • The proposed method was used for obtaining monodisperse and uniform silver nanoparticles-loaded chitosan microparticles measuring numerous hundred micrometers. • In addition, together with the benefits of flexible manipulation and largescale integration, microfluidic technology has been hastily developed as one of the most significant platforms in the field of purposeful biomaterial synthesis.
 ADVANTAGE :
 ? There is no guarantee that the amplification efficiency will remain constant during the reaction. ? However, there are many factors that affect the amplification efficiency during PCR amplification. ? In other words, the amplification efficiency may be different between the work sample and the standard sample. ? A complementary metal oxide semiconductor (CMOS) camera was used as a detection tool to obtain fluorescence video for the entire loop area or a specified loop area. ? With the development of life science and medical testing, it has become increasingly important that faster and more accurate disease detection devices can be used for more people.

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