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Molecular Analysis in Mechanobiology
Published in Jiro Nagatomi, Eno Essien Ebong, Mechanobiology Handbook, 2018
PCR was invented by Dr. Kary Mullis and colleagues at Cetus Corp in the mid-1980s [26,27]. The reaction consists of a template DNA molecule, two short DNA oligonucleotides complimentary to the template (primers), DNA polymerase, buffer with appropriate Mg2+ concentration, and dNTPs. Through a process of thermal cycling, a specific region of the DNA template is selectively amplified. The use of a thermostable DNA polymerase (Taq polymerase) derived from the bacterium Thermus aquaticus substantially improved the efficiency and utility of PCR [28]. Reverse transcriptase, an RNA-dependent DNA polymerase that synthesizes cDNA from an RNA template allowed the application of PCR to mRNA analysis (RT-PCR).
Enzymes Used for Recombinant DMA Technology Produced by Recombinant Microbes
Published in Yoshikatsu Murooka, Tadayuki Imanaka, Recombinant Microbes for Industrial and Agricultural Applications, 2020
Various types of DNA polymerases are also used in recombinant DNA technology, especially for DNA sequencing and polymerase chain reaction (PCR). Most of these enzymes require a template and synthesize a product the sequence of which is complementary to that of the template. The DNA-dependent DNA polymerases used frequently are E. coli DNA polymerase I, large fragment of E. coli DNA polymerase I (Klenow fragment), modified T7 DNA polymerase, and Taq DNA polymerase. A reverse transcriptase (RNA-dependent DNA polymerase) prefers to copy RNA. Properties of these DNA polymerases are summarized in Table 3. All of these DNA polymerases are now purified from recombinant microbes.
Detection Technology
Published in Rick Houghton, William Bennett, Emergency Characterization of Unknown Materials, 2020
Rick Houghton, William Bennett
Polymerase chain reaction technology uses a polymerase, a naturally occurring enzyme, to catalyze the formation of DNA or a characteristic segment of DNA. DNA is formed from two complementary, not identical, chains of amino acid, which are bound together. If the DNA is torn in half (down the center, like a zipper), polymerase can recreate the complimentary chain missing from each half, which results in two strands of DNA identical to the beginning chain (Figure 3.54).
Cost-effective, high-yield production of Pyrobaculum calidifontis DNA polymerase for PCR application
Published in Preparative Biochemistry & Biotechnology, 2023
Kashif Maseh, Syed Farhat Ali, Shazeel Ahmad, Naeem Rashid
DNA polymerase is an important enzyme used for various applications and processes including routine PCR, real-time PCR, error-prone PCR, site-directed mutagenesis, DNA labeling and sequencing.[12,16] In addition, DNA polymerases are used for applications in synthetic biology and DNA-based computing.[17,18] Here we report gene expression optimization and high-recovery affinity purification of Pca-Pol. This enzyme has been shown to amplify DNA fragments up to 7.5 kb and used for DNA labeling application as well.[14]