Polymerase amplification

Gene analysis DNA polymerase Amplification of gene sequences by PCR and electrophoretic separation of PCR products Fluorescence... 

PGR amplification of a DNA sequence is faciHtated by the use of a heat-stable DNA polymerase, Taq polymerase (TM), derived from the thermostable bacterium Thermus aquaticus. The thermostable polymerase allows the repeated steps of strand separation, primer annealing, and DNA synthesis to be carried out ia a single reactioa mixture where the temperature is cycled automatically. Each cycle coasists of a high temperature step to deaature the template strands, a lower temperature annealing of the primer and template, and a higher temperature synthesis step. AH components of the reaction are present ia the same tube. 

FIGURE 28.14 The polymerase chain reaction (PCR). Three cycles are shown the target region appears after the third cycle. Additional cycles lead to amplification of the target region. 

Kasai K, Nakamura Y and White R 1990) Amplification of a variable number of tandem repeats (VNTR) locus (pMCTii8) by the polymerase chain reaction (PCR) and its application to forensic science. ] For Sci 35 1196-1200. 

C. E. Gustafson, C. J. Thomas, and T, J. Trust, Detection of Aera/tm/tav-salmoni-cida from fish by using polymerase chain-reaction amplification of the virulence surface array protein gene. Appl. Environ. Microbiol., 5S 3825 (1992). 

The linear RCA method can use both target and signal amplification. A DNA circle (such as a plasmid, circular vims or circular chromosome) is amplified by polymerase extension of a complementary primer. Up to 10 tandemly repeated, concantemerized copies of the DNA circle are generated by each primer, resulting in one single-stranded, concantemerized product. ... 

Synthetic oligonucleotides are very important tools in the study and manipulation of DNA, including such techniques as site-directed mutagenesis and DNA amplification by the polymerase chain reaction. The techniques for chemical synthesis of oligonucleotides are highly developed. Very efficient automated methodologies based on solid phase synthesis are used extensively in fields that depend on the availability of defined DNA sequences.52... 

Polymerase chain reaction (PCR DNA amplification of the most common bacterial meningitis pathogens) may be useful to help exclude bacterial meningitis. 

Molecular methods used to uncover mutations are subject to several variables. The anticoagulants used for blood collection can affect digestion with restriction enzymes and amplification reactions. The type of detergent used in cell lysis can affect amplification of DNA by inhibiting the DNA-amplifying enzyme such as the taq polymerase used in the polymerase chain reaction (116). The control of contamination is crucial in ensuring the quality of results obtained by molecular analysis (117). 

Both target and signal amplification systems have been successfully employed to detect and quantitate specific nucleic acid sequences in clinical specimens. Polymerase chain reaction (PCR), nucleic acid sequence-based amplification (NASBA), transcription-mediated amplification (TMA), strand displacement amplification (SDA), and ligase chain reaction (LCR) are all examples of enzyme-mediated, target amplification strategies that are capable of producing billions of... 

Saiki, R. K., etal. (1988). Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science 239,487-491. 

Iqbal, S. Robinson, J. Deere, D. Saunders, J. R. Edwards, C. Porter, J. Efficiency of the polymerase chain reaction amplification of the uid gene for detection of Escherichia coli in contaminated water. Lett. Appl. Microbiol. 1997,24,498-502. 

Mullis, K.B., Faloona, F., Scharf, S., Saiki, R., Horn, G. and Erlich, H. (1986) Specific enzymatic amplification of DNA in vitro the polymerase chain reaction. Cold Spring Harbor Symposium of Quantitative Biology 51, 263—273. 

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