DNA polymerase

Real-time PCR fluorescence detection systems

Real-time PCR fluorescence detection systems Real-time fluorescent PCR chemistries Many real-time fluorescent PCR chemistries exist, but the most widely used are 5” nuclease assays such as TaqMan®Assays and SYBR® Green dye–based assays (Figure 1). The 5” nuclease assay is named for the 5” nuclease activityassociated with Taq DNA polymerase (Figure 2). The 5” nuclease domain […]

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REAL-TIME PCR COMPONENTS

DNA polymerasePCR performance is often related to the thermostable DNA polymerase, so enzyme selection is critical to success. One of the main factors affecting PCR specificity is the fact that Taq DNA polymerase has residual activity at low temperatures. Primers can anneal nonspecifically to DNA during reaction setup, allowing the polymerase to synthesize nonspecific product.

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STEPS INVOLVED IN PERFORMING REAL-TIME PCR

Real-time PCR is a variation of the standard PCR technique that is commonly used to quantify DNA or RNA in a sample. Using sequence-specific primers, the number of copies of a particular DNA or RNA sequence can be determined. By measuring the amount of amplified product at each stage during the PCR cycle, quantification is

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HOW TO DESIGN PRIMERS FOR YOUR PCR EXPERIMENT

Primers are short stretches of DNA that target unique sequences of a DNA molecule and help identify a unique part of a genome or a gene to be copied further. They are usually 18 to 25 nucleotides long. Primers can be synthesized and used for your PCR (polymerase chain reaction) and other molecular biology experiment

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