PCR

PCR

PCR Animation

If the nucleotide sequences at the ends of a particular DNA region are known, the intervening fragment can be amplified directly by the polymerase chain reaction (PCR). Here we describe the basic PCR technique and three situations in which it is used.

The PCR depends on the ability to alternately denature (melt) double-stranded DNA molecules and renature (anneal) complementary single strands in a controlled fashion. As in the membrane-hybridization assay described earlier, the presence of noncomplementary strands in a mixture has little effect on the base pairing of complementary single DNA strands or complementary regions of strands. The second requirement for PCR is the ability to synthesize oligonucleotides at least 18–20 nucleotides long with a defined sequence. Such synthetic nucleotides can be readily produced with automated instruments based on the standard reaction scheme.

A typical PCR procedure begins by heat-denaturation of a DNA sample into single strands. Next, two synthetic oligonucleotides complementary to the 3' ends of the target DNA segment of interest are added in great excess to the denatured DNA, and the temperature is lowered to 50–60 C. These specific oligonucleotides, which are at a very high concentration, will hybridize with their complementary sequences in the DNA sample, whereas the long strands of the sample DNA remain apart because of their low concentration. The hybridized oligonucleotides then serve as primers for DNA chain synthesis in the presence of deoxynucleotides (dNTPs) and a temperature-resistant DNA polymerase such as that from Thermus aquaticus (a bacterium that lives in hot springs).

This enzyme, called Taq polymerase, can remain active even after being heated to 95 C and can extend the primers at temperatures up to 72 C. When synthesis is complete, the whole mixture is then heated to 95 C to melt the newly formed DNA duplexes. After the temperature is lowered again, another cycle of synthesis takes place because excess primer is still present. Repeated cycles of melting (heating) and synthesis (cooling) quickly amplify the sequence of interest. At each cycle, the number of copies of the sequence between the primer sites is doubled; therefore, the desired sequence increases exponentially—about a million-fold after 20 cycles—whereas all other sequences in the original DNA sample remain unamplified.

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  • Ureaplasma parvum/Ur.urealyticum/ M.hominis RT-PCR Quant (CE) | B75-FRT Q

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  • Shigella/Salmonella/Campylobacter RT-PCR | B44-FRT

    Shigella/Salmonella/Campylobacter RT-PCR (CE) | B44-FRT

    Select Currency //740.00 - Select Currency //950.00
    Shigella/Salmonella/Campylobacter RT-PCR | B44-FRT from Sacace Biotechnologies is available for delivery Real Time PCR Kit for detection and differentiation of Shigella spp and Enteroinvasive E.Coli (EIEC)/ Salmonella spp/Campylobacter spp Storage &...
    441-B44-FRT
    Select Currency //740.00 - Select Currency //950.00
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  • Borrelia burgdorferi RT-PCR | B37-FRT

    Borrelia burgdorferi RT-PCR (CE) | B37-FRT

    Select Currency //740.00 - Select Currency //950.00
    Borrelia burgdorferi RT-PCR | B37-FRT from Sacace Biotechnologies is available for delivery Real Time PCR Kit for qualitative detection of Borrelia burgdorferi in biological materials Storage & Shipping : The kit can be shipped at 2-8°C for 3-4...
    441-B37-FRT
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  • Listeria monocytogenes RT-PCR Quant | B14-FRT

    Listeria monocytogenes RT-PCR Quant (CE) | B14-FRT

    Select Currency //740.00 - Select Currency //950.00
    Listeria monocytogenes RT-PCR Quant | B14-FRT from Sacace Biotechnologies is available for delivery Real Time PCR Kit for quantitative detection of Listeria monocytogenes Storage & Shipping : The kit can be shipped at 2-8°C for 3-4 days but...
    441-B14-FRT
    Select Currency //740.00 - Select Currency //950.00
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  • Shigella spp & EIEC RT-PCR | B12-FRT

    Shigella spp & EIEC RT-PCR (CE) | B12-FRT

    Select Currency //740.00 - Select Currency //950.00
    Shigella spp & EIEC RT-PCR | B12-FRT from Sacace Biotechnologies is available for delivery Real Time PCR Kit for detection of Shigella spp andEnteroinvasive E.Coli (EIEC) Storage & Shipping : The kit can be shipped at 2-8°C for 3-4 days but...
    441-B12-FRT
    Select Currency //740.00 - Select Currency //950.00
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  • Helicobacter pylori RT-PCR | B9-FR

    Helicobacter pylori RT-PCR (CE) | B9-FR

    Select Currency //740.00 - Select Currency //950.00
    Helicobacter pylori RT-PCR | B9-FR from Sacace Biotechnologies is available for delivery Real Time PCR kit for qualitative detection of Helicobacter pylori Storage & Shipping :The kit can be shipped at 2-8°C for 3-4 days but should be stored at 2-8°C and...
    441-B9-FR
    Select Currency //740.00 - Select Currency //950.00
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