Polymerase Chain Reaction And Its Applications Pdf

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David N. Fredricks, David A. Oxford University Press is a department of the University of Oxford.

Polymerase Chain Reaction (PCR) Fact Sheet

Sometimes called "molecular photocopying," the polymerase chain reaction PCR is a fast and inexpensive technique used to "amplify" - copy - small segments of DNA. Because significant amounts of a sample of DNA are necessary for molecular and genetic analyses, studies of isolated pieces of DNA are nearly impossible without PCR amplification. Often heralded as one of the most important scientific advances in molecular biology, PCR revolutionized the study of DNA to such an extent that its creator, Kary B. Mullis, was awarded the Nobel Prize for Chemistry in PCR is also valuable in a number of laboratory and clinical techniques, including DNA fingerprinting, detection of bacteria or viruses particularly AIDS , and diagnosis of genetic disorders.

The polymerase chain reaction PCR is a powerful research tool used in many scientific disciplines. It is also used for detection and testing in areas such as food microbiology, environmental microbiology, biotechnology, industrial microbiology, veterinary and medical diagnostics. This indispensable manual is a compilation of review articles written by experts in the field of PCR technology. Topics covered include: principles of PCR, fluorescent chemistries, instrumentation, quantification strategies, extraction and purification of nucleic acids, sample preparation, controls for validation, primers and probes, standardization of methods, MIQE guidelines, mRNA expression and PCR arrays. This book provides a comprehensive overview of PCR theory, instrumentation and methods. The book represents an excellent, detailed guide for anyone interested in the development and use of PCR technology. It is a recommended purchase for all microbiology and molecular biology laboratories and university libraries.

Real-time polymerase chain reaction

Metrics details. Based upon defining a common reference point, current real-time quantitative PCR technologies compare relative differences in amplification profile position. As such, absolute quantification requires construction of target-specific standard curves that are highly resource intensive and prone to introducing quantitative errors. Sigmoidal modeling using nonlinear regression has previously demonstrated that absolute quantification can be accomplished without standard curves; however, quantitative errors caused by distortions within the plateau phase have impeded effective implementation of this alternative approach. Recognition that amplification rate is linearly correlated to amplicon quantity led to the derivation of two sigmoid functions that allow target quantification via linear regression analysis. In addition to circumventing quantitative errors produced by plateau distortions, this approach allows the amplification efficiency within individual amplification reactions to be determined. Absolute quantification is accomplished by first converting individual fluorescence readings into target quantity expressed in fluorescence units, followed by conversion into the number of target molecules via optical calibration.

International Journal of Tropical Medicine

Principles and applications of polymerase chain reaction in medical diagnostic fields: a review. Recent developments in molecular methods have revolutionized the detection and characterization of microorganisms in a broad range of medical diagnostic fields, including virology, mycology, parasitology, microbiology and dentistry. PCR is an excellent technique for the rapid detection of pathogens, including those difficult to culture.

Two common methods for the detection of PCR products in real-time PCR are 1 non-specific fluorescent dyes that intercalate with any double-stranded DNA and 2 sequence-specific DNA probes consisting of oligonucleotides that are labelled with a fluorescent reporter, which permits detection only after hybridization of the probe with its complementary sequence. Cells in all organisms regulate gene expression by turnover of gene transcripts single stranded RNA : The amount of an expressed gene in a cell can be measured by the number of copies of an RNA transcript of that gene present in a sample. In order to robustly detect and quantify gene expression from small amounts of RNA, amplification of the gene transcript is necessary. In order to amplify small amounts of DNA, the same methodology is used as in conventional PCR using a DNA template, at least one pair of specific primers , deoxyribonucleotides , a suitable buffer solution and a thermo-stable DNA polymerase.

Arch Neurol. Polymerase chain reaction PCR is a broadly applied laboratory test for the diagnosis of a wide variety of central nervous system CNS diseases, including genetic and autoimmune diseases, malignant neoplasms, and infections. In addition, PCR can be performed on a variety of tissues preserved in different ways—even archival specimens can be used to provide important epidemiological information. By making quick and precise diagnoses, appropriate treatments can be instituted, and unnecessary or invasive investigations can be avoided. The power of PCR results from its ability to synthesize millions of copies of a specific gene segment in vitro, starting with one or only a few template copies, an amount undetectable by other methods.

2015, Number 3

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