Bead-based polymerase chain reaction on a microchip

We present a bead-based approach to microfluidic polymerase chain reaction (PCR), enabling fluorescent detection and sample conditioning in a single microchamber. Bead-based PCR, while not extensively investigated in microchip format, has been used in a variety of bioanalytical applications in recent years. We leverage the ability of bead-based PCR to accumulate fluorescent labels following DNA amplification to explore a novel DNA detection scheme on a microchip. The microchip uses an integrated microheater and temperature sensor for rapid control of thermal cycling temperatures, while the sample is held in a microchamber fabricated from (poly)dimethylsiloxane and coated with Parylene. The effects of key bead-based PCR parameters, including annealing temperature and concentration of microbeads in the reaction mixture, are studied to achieve optimized device sensitivity and detection time. The device is capable of detecting a synthetically prepared section of the Bordetella pertussis genome in as few as 10 temperature cycles with times as short as 15?min. We then demonstrate the use of the procedure in an integrated device; capturing, amplifying, detecting, and purifying template DNA in a single microfluidic chamber. These results show that this method is an effective method of DNA detection which is easily integrated in a microfluidic device to perform additional steps such as sample pre-conditioning.     

Continuous flow polymerase chain reaction using a hybrid PMMA-PC microchip with improved heat tolerance

Recently, polymeric materials have been explored as more versatile alternatives for the fabrication of polymerase chain reaction (PCR) microchips. Poly(methyl methacrylate) (PMMA) is a popular substrate material due to its high mechanical stability, good chemical properties and most importantly, its suitability for cheap and simple CO₂ laser ablation. However, it has a low glass transition temperature (T_g) of 105 °C, which is just above the denaturation temperature for PCR, thus the bond integrity is compromised. Polycarbonate (PC) is preferred as a substrate for PCR microchip as it has a higher T_g of 150 °C; but since its thermal properties are not suitable for CO₂ laser light, the more expensive excimer laser has to be employed. Here we report a novel hybrid PMMA-PC microchip by bonding a PC cover plate with a PMMA substrate containing microchannel which is fabricated by CO₂ laser ablation. This hybrid microchip has improved heat tolerance, such that the bonding integrity is sustained at the denaturation temperature. DNA amplification is found to be more efficiently performed in a PMMA-PC microchip than in a PMMA-PMMA microchip.     

The design and fabrication of polymerase chain reaction platform

Thermalcycler was extensively used machine for amplify DNA sample. One of the major problems in the working time was that it spent most of time for cooling and heating. In order to improve the efficient, the study presented a novel method for amplify DNA sample. For this concept, the DNA sample in the silicon chamber which was pushed by a tappet through the three temperature regions around a center and then the DNA segments could be amplified rapidly after 30 cycles. The polymerase chain reaction (PCR) platform was composed of the thin-film heaters, Cu plates, DC powers, and temperature controllers. The photolithography and bulk etching technologies were utilized to construct the thin-film heater and DNA reaction chambers. Finally, 1 μl 100 bp DNA segment of Escherichia coli K12 was amplified successfully within 36 min on this PCR platform.     

聚合酶链式反应微流控生物芯片的数值模拟

微通道内具有一定流速的DNA反应混和物能否达到聚合酶链式反应(PCR)指定的温度PCR微流控芯片研究的关键问题.本文应用有限体元法(FEV)数值模拟该芯片上3个恒温区的直型、弯型、逶迤型三类微通道内,微流体的温度场和速度场.结果表明:对于宽100 μm深50μm的微通道,速度在0.002 m/s～0.02 m/s范围内,180.的弯道以及温度场、温度梯度的存在对其速度场分布无影响,微流体仍旱现为层流;微流体大约经过60μm的距离,其温度场达到稳态,其速度场充分发展为层流;采用宽4 mm深2 mm的空气隔热槽能起到隔热的效果.     

A new thermal cycling mechanism for effective polymerase chain reaction in microliter volumes

This study presents a new thermal cycler using infrared (IR) heating and water impingement cooling for polymerase chain reaction (PCR) amplification of 10 l samples in thin glass capillary tubes. The thermal cycling system can achieve a temperature ramping-up rate of 65 °C/s and a ramping-down rate of 80 °C/s. Two other cooling mechanisms, natural convection and forced air convection, can also be used in the present system to obtain a ramping-down rate of 2 °C/s and 6 °C/s, respectively. The amplification of the 439 fragment of hepatitis B virus (HBV) DNA was successful. The PCR amplified products were analyzed by agarose gel electrophoresis with ethidium bromide staining for visualization. A comparison of results of the amplification products at three different ramping-down rates was made and the rapid thermal cycling of the present system can run DNA required amplification in 29 min for 20 thermal cycles that is only 1/3 the time spent in the conventional PCR machine used in comparison.This work was supported by the National Taiwan University College of Medicine in preparing the HBV DNA samples. We thank our research assistance W. L. Chen and Dr. Chen for their technical advice. We gratefully acknowledge the financial support by the Nation Science Council, No. NSC-88–2212-E-002–025.     

On-chip polymerase chain reaction microdevice employing a magnetic droplet-manipulation system

An on-chip polymerase chain reaction (PCR) device employing a magnetic beads-droplet-handling system was developed. Actuation with a magnet offers a simple system for droplet manipulation that allows separation and fusion of droplets containing magnetic beads by handling with a magnet. The device consists of a reaction chamber channel and two magnet-handling channels for the manipulation of micro-droplets containing magnetic beads. Micro-droplets were placed inside a reaction chamber filled with oil and manipulated with a magnet. When a droplet containing NaOH and magnetic beads was manipulated towards a droplet containing phenol red, a color change was observed after fusion. Sample preparation was performed by fusion of droplets containing a forward primer, reverse primer, template DNA and PCR mixture, using a droplet containing magnetic beads. PCR amplification or RT-PCR was also successfully performed, with efficiency comparable to manual methods that use this device by placing it on a thermal cycler for amplification. With a magnetic beads-manipulation step, purification of amplified DNA was also accomplished by using magnetic beads as the carrier. The amplified DNA was captured on streptavidin conjugated magnetic beads using a biotinylated primer, purified by washing and digested for separation of the target DNA.     

Enhancement of thermal uniformity for a microthermal cycler and its application for polymerase chain reaction

Thermal uniformity is essentially important for micro reactors which require precise control of critical reaction temperatures. Accordingly, we report a new approach to increase the temperature uniformity inside a microthermal cycler, especially for polymerase chain reaction (PCR). It enhances the thermal uniformity in the reaction region of a PCR chip by using new array-type microheaters with active compensation (AC) units. With this approach, the edges of the microthermal cyclers which commonly have significant temperature gradients can be compensated. Significantly, the array-type microheaters provide higher uniformity than conventional block-type microheaters. Besides, experimental data from infrared (IR) images show that the percentages of the uniformity area with a thermal variation of less than 1 °C are 63.6%, 96.6% and 79.6% for three PCR operating temperatures (94, 57 and 72 °C, respectively) for the new microheaters. These values are significantly better than the conventional block-type microheaters. Finally, the performance of this proposed microthermal cycler is successfully demonstrated by amplifying a detection gene associated with Streptococcus Pneumoniae (S. Pneumoniae). The PCR efficiency of the new microthermal cycler is statistically higher than the block-type microheaters. Therefore, the proposed microthermal cycler is suitable for DNA amplification which requires a high temperature uniformity and is crucial for micro reactors with critical thermal constraints.     

Comparison of different methods to analyze a DNA computing library using the polymerase chain reaction

Screening and analysis of collections of DNA molecules is a standard aspect of many DNA computing approaches. We describe the use of three different polymerase chain reaction (PCR) detection methods to screen specific members of a 5-site, 2-variable DNA computing library previously created using parallel overlap assembly of unique sequences generated from the SynDCode program. The three PCR methods (conventional gel-based PCR, SYBR Green real-time PCR and TaqMan real-time PCR) could all successfully identify individual library members separately or in a mixture. The TaqMan approach was also able to identify members in the original library we had not yet sequenced, providing more evidence supporting our hypothesis that the DNA library we generated may be complete. We expect these three approaches will be useful in future screening of other DNA computing libraries and structures.     

基于神经网络的注塑机注射速度的迭代学习控制

对具有不确定性和干扰项的重复非线性注塑机控制系统,尤其是注射速度的控制,提出基于神经网络的迭代学习控制器,其中迭代学习控制器设计为神经网络控制器,它以前馈方式作用于对象。PD反馈控制器用于使系统达到稳定,同时和前馈的神经网络学习控制器一起使系统达到理想的控制效果。仿真结果表明,该控制器可以随着迭代次数的增加有效减小跟踪误差。     

Adaptive backstepping control of wheeled inverted pendulum with velocity estimator

In this paper, we introduce a backstepping control design of a wheeled inverted pendulum. Based on a second-order motion equation of the body angle, an adaptive integral backstepping controller is designed to stabilize the body angle. It is shown that the σ-modification rule in the adaptive update law guarantees the boundedness of the errors in estimating the time-varying signal that is an output of a linear system with every bounded input signal. Then, the stabilizing controller for the wheel angle is constructed by a PD-type positive feedback. The derived controller requires the full-state measurements. In the output feedback case, the K filter or the observer backstepping is needed. However, the structure of the controller becomes complicated. We propose a non-model-based differentiator based on the adaptive update law. Since the non-model-based differentiator does not require any knowledge of the dynamic structure of the signal, we can use it as a velocity estimator for unknown nonlinear systems. Therefore, we replaced the velocity measurement with the estimates by the non-model-based differentiator. Finally, simulation results for the proposed controller are presented.     

An improved approach to H-two control with regional stability constraints

This paper presents an improved method for H-two control with regional stability constraints for the closedloop system. The controller is given by the convex combination of a set of fixed gains. Both continuous and discrete-time systems in a known polytopic domain are investigated. New LMI-based sufficient conditions for the existence of parameterdependent Lyapunov functions are proposed. Numerical examples are given to show the proposed conditions provide useful and less conservative results for the problems of H-two control with regional stability constraints.     

A class of time-optimal velocity control laws for a spinning space vehicle

Several time-optimal velocity control laws are described for a spinning space vehicle. It is shown that several laws, rather than a unique law, result from the non-normality of the system. Given initial pitch and yaw body rates, a control system is defined that will drive these rates to zero in minimum time.     

A concept of the terminal descent control algorithm at reentry into the earth atmosphere with near-parabolic velocity

A concept of the terminal descent control algorithm of the descent module at reentry into the Earth atmosphere with near-parabolic velocity is proposed. Trajectories with one and two (skip reentry) reentries are considered. Simultaneously, downrange and crossrange misses are eliminated by controlling the bank angle and time of bank angle reversal at each phase of the trajectory. The designed control meets the specified load factor constraint.     

Robust adaptive motion control for underwater remotely operated vehicles with velocity constraints

In this paper, robust adaptive control strategies are designed for Underwater Remotely Operated Vehicles (ROVs) with velocity constraints. First, robust control strategies are investigated for under-water ROVs, and then adaptive robust control strategies are further developed with online parameter estimation. To prevent the velocity constraint violation, the Barrier Lyapunov Function (BLF) is employed in Lyapunov synthesis. By ensuring the boundedness of the BLF, we also guarantee that the velocity constraints are not transgressed. The stability analysis of the closed-loop system is provided and all closed-loop signals are ensured to be bounded. Simulation results for 5 degree-of-freedom (DOF) underwater ROV demonstrate the effectiveness of the proposed approach.     

Dynamic modeling and control of supply chain systems: A review

Supply chains are complicated dynamical systems triggered by customer demands. Proper selection of equipment, machinery, buildings and transportation fleets is a key component for the success of such systems. However, efficiency of supply chains mostly depends on management decisions, which are often based on intuition and experience. Due to the increasing complexity of supply chain systems (which is the result of changes in customer preferences, the globalization of the economy and the stringy competition among companies), these decisions are often far from optimum. Another factor that causes difficulties in decision making is that different stages in supply chains are often supervised by different groups of people with different managing philosophies. From the early 1950s it became evident that a rigorous framework for analyzing the dynamics of supply chains and taking proper decisions could improve substantially the performance of the systems. Due to the resemblance of supply chains to engineering dynamical systems, control theory has provided a solid background for building such a framework. During the last half century many mathematical tools emerging from the control literature have been applied to the supply chain management problem. These tools vary from classical transfer function analysis to highly sophisticated control methodologies, such as model predictive control (MPC) and neuro-dynamic programming. The aim of this paper is to provide a review of this effort. The reader will find representative references of many alternative control philosophies and identify the advantages, weaknesses and complexities of each one. The bottom line of this review is that a joint co-operation between control experts and supply chain managers has the potential to introduce more realism to the dynamical models and develop improved supply chain management policies.     

A Hough Transform with projection for velocity estimation

An extended Hough Transform is proposed to estimate the velocity of Regions Of Interest (ROI). Each ROI is first characterized by a spatio-temporal point (centroids for example). The velocity of each point is then computed by searching the line that passes through this point and fits at best the other ones. In order to avoid high dimension problems (a three-dimensional (3D) spatio-temporal line is defined by four parameters), the data are projected on several planes. The velocities are deduced by combining the results obtained on each plane using weighted least squares. The robustness of this method called “Hough Transform with Projection (HTP)”, compared to the Standard Hough Transform (applied on 3D spatio-temporal data) is demonstrated. Finally, experimental results on synthetic and real sequences validate the approach.