Commenced in January 2007
Frequency: Monthly
Edition: International
Paper Count: 6

Polymerase chain reaction Related Publications

6 Negative RT-PCR in a Newborn Infected with Zika Virus: A Case Report

Authors: Vallejo Michael, Acuña Edgar, Roa Juan David, Peñuela Rosa, Parra Alejandra, Casallas Daniela, Rodriguez Sheyla

Abstract:

Congenital Zika Virus Syndrome is an entity composed by a variety of birth defects presented in newborns that have been exposed to the Zika Virus during pregnancy. The syndrome characteristic features are severe microcephaly, cerebral tissue abnormalities, ophthalmological abnormalities such as uveitis and chorioretinitis, arthrogryposis, clubfoot deformity and muscular tone abnormalities. The confirmatory test is the Reverse transcription polymerase chain reaction (RT-PCR) associated to the physical findings. Here we present the case of a newborn with microcephaly whose mother presented a confirmed Zika Virus infection during the third trimester of pregnancy, despite of the evident findings and the history of Zika infection the RT-PCR in amniotic and cerebrospinal fluid of the newborn was negative. RT-PCR has demonstrated a low sensibility in samples with low viral loads, reason why, we propose a clinical diagnosis in patients with clinical history of Zika Virus infection during pregnancy accompanied by evident clinical manifestations of the child.

Keywords: Polymerase chain reaction, Zika virus, microcephaly, amniotic fluid

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5 PTFE Capillary-Based DNA Amplification within an Oscillatory Thermal Cycling Device

Authors: Jyh J. Chen, Fu H. Yang, Ming H. Liao

Abstract:

This study describes a capillary-based device integrated with the heating and cooling modules for polymerase chain reaction (PCR). The device consists of the reaction polytetrafluoroethylene (PTFE) capillary, the aluminum blocks, and is equipped with two cartridge heaters, a thermoelectric (TE) cooler, a fan, and some thermocouples for temperature control. The cartridge heaters are placed into the heating blocks and maintained at two different temperatures to achieve the denaturation and the extension step. Some thermocouples inserted into the capillary are used to obtain the transient temperature profiles of the reaction sample during thermal cycles. A 483-bp DNA template is amplified successfully in the designed system and the traditional thermal cycler. This work should be interesting to persons involved in the high-temperature based reactions and genomics or cell analysis.

Keywords: Capillary, Polymerase chain reaction, thermal cycles, TE cooler

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4 Design and Microfabrication of a High Throughput Thermal Cycling Platform with Various Annealing Temperatures

Authors: Sin J. Chen, Jyh J. Chen

Abstract:

This study describes a micro device integrated with multi-chamber for polymerase chain reaction (PCR) with different annealing temperatures. The device consists of the reaction polydimethylsiloxane (PDMS) chip, a cover glass chip, and is equipped with cartridge heaters, fans, and thermocouples for temperature control. In this prototype, commercial software is utilized to determine the geometric and operational parameters those are responsible for creating the denaturation, annealing, and extension temperatures within the chip. Two cartridge heaters are placed at two sides of the chip and maintained at two different temperatures to achieve a thermal gradient on the chip during the annealing step. The temperatures on the chip surface are measured via an infrared imager. Some thermocouples inserted into the reaction chambers are used to obtain the transient temperature profiles of the reaction chambers during several thermal cycles. The experimental temperatures compared to the simulated results show a similar trend. This work should be interesting to persons involved in the high-temperature based reactions and genomics or cell analysis.

Keywords: Polymerase chain reaction, Micro-Fabrication, thermal cycles, temperature gradient

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3 Mutational Analysis of CTLA4 Gene in Pakistani SLE Patients

Authors: N. Hussain, G. Jaffery, A.N. Sabri, S. Hasnain

Abstract:

The main aim is to perform mutational analysis of CTLA4 gene Exon 1 in SLE patients. A total of 61 SLE patients fulfilling “American College of Rheumatology (ACR) criteria" and 61 controls were enrolled in this study. The region of CTLA4 gene exon 1 was amplified by using Step-down PCR technique. Extracted DNA of band 354 bp was sequenced to analyze mutations in the exon-1 of CTLA-4 gene. Further, protein sequences were identified from nucleotide sequences of CTLA4 Exon 1 by using Expasy software and through Blast P software it was found that CTLA4 protein sequences of Pakistani SLE patients were similar to that of Chinese SLE population. No variations were found after patients sequences were compared with that of the control sequence. Furthermore it was found that CTLA4 protein sequences of Pakistani SLE patients were similar to that of Chinese SLE population. Thus CTLA4 gene may not be responsible for an autoimmune disease SLE.

Keywords: Autoimmune Disease, systemic lupus erythematosus, Polymerase chain reaction, American College of Rheumatology criteria, Cytotoxic T Lymphocyte Antigen-4

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2 Simulation and Design of the Geometric Characteristics of the Oscillatory Thermal Cycler

Authors: Tse-Yu Hsieh, Jyh-Jian Chen

Abstract:

Since polymerase chain reaction (PCR) has been invented, it has emerged as a powerful tool in genetic analysis. The PCR products are closely linked with thermal cycles. Therefore, to reduce the reaction time and make temperature distribution uniform in the reaction chamber, a novel oscillatory thermal cycler is designed. The sample is placed in a fixed chamber, and three constant isothermal zones are established and lined in the system. The sample is oscillated and contacted with three different isothermal zones to complete thermal cycles. This study presents the design of the geometric characteristics of the chamber. The commercial software CFD-ACE+TM is utilized to investigate the influences of various materials, heating times, chamber volumes, and moving speed of the chamber on the temperature distributions inside the chamber. The chamber moves at a specific velocity and the boundary conditions with time variations are related to the moving speed. Whereas the chamber moves, the boundary is specified at the conditions of the convection or the uniform temperature. The user subroutines compiled by the FORTRAN language are used to make the numerical results realistically. Results show that the reaction chamber with a rectangular prism is heated on six faces; the effects of various moving speeds of the chamber on the temperature distributions are examined. Regarding to the temperature profiles and the standard deviation of the temperature at the Y-cut cross section, the non-uniform temperature inside chamber is found as the moving speed is larger than 0.01 m/s. By reducing the heating faces to four, the standard deviation of the temperature of the reaction chamber is under 1.4×10-3K with the range of velocities between 0.0001 m/s and 1 m/s. The nature convective boundary conditions are set at all boundaries while the chamber moves between two heaters, the effects of various moving velocities of the chamber on the temperature distributions are negligible at the assigned time duration.

Keywords: Polymerase chain reaction, oscillatory thermal cycler, standard deviation of temperature, nature convective

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1 The Association of Matrix Metalloproteinase-3 Gene -1612 5A/6A Polymorphism with Susceptibility to Coronary Artery Stenosis in an Iranian Population

Authors: M. Seifi, S. Fallah, M. Firoozrai

Abstract:

Matrix metalloproteinase-3 (MMP3) is key member of the MMP family, and is known to be present in coronary atherosclerotic. Several studies have demonstrated that MMP-3 5A/6A polymorphism modify each transcriptional activity in allele specific manner. We hypothesized that this polymorphism may play a role as risk factor for development of coronary stenosis. The aim of our study was to estimate MMP-3 (5A/6A) gene polymorphism on interindividual variability in risk for coronary stenosis in an Iranian population.DNA was extracted from white blood cells and genotypes were obtained from coronary stenosis cases (n=95) and controls (n=100) by PCR (polymerase chain reaction) and restriction fragment length polymorphism techniques. Significant differences between cases and controls were observed for MMP3 genotype frequencies (X2=199.305, p< 0.001); the 6A allele was less frequently seen in the control group, compared to the disease group (85.79 vs. 78%, 6A/6A+5A/6A vs. 5A/5A, P≤0.001). These data imply the involvement of -1612 5A/6A polymorphism in coronary stenosis, and suggest that probably the 6A/6A MMP-3 genotype is a genetic susceptibility factor for coronary stenosis.

Keywords: Polymerase chain reaction, Polymorphism, Coronary artery stenosis, matrixmetalloproteinase-3

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