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Mechanism-based qPCR quantification methods have also been suggested, and have the advantage that they do not require a standard curve for quantification. Methods such as MAK2 [16] have been shown to have equal or better quantitative performance to standard curve methods. These mechanism-based methods use knowledge about the polymerase ...
The Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines are a set of protocols for conducting and reporting quantitative real-time PCR experiments and data, as devised by Bustin et al. in 2009. [1]
Chimeric polymerases overcome many limitations of native enzymes and are used in direct PCR amplification from cell cultures and even food samples, thus by-passing laborious DNA isolation steps. A robust strand-displacement activity of the hybrid TopoTaq polymerase helps solve PCR problems that can be caused by hairpins and G-loaded double helices.
Some modifications are needed to perform long PCR. The original Klenow-based PCR process did not generate products that were larger than about 400 bp. Taq polymerase can however amplify targets of up to several thousand bp long. [4] Since then, modified protocols with Taq enzyme have allowed targets of over 50 kb to be amplified. [5]
Besides reporting guidelines, the MIQE stresses the need to standardize the nomenclature associated with quantitative PCR to avoid confusion; for example, the abbreviation qPCR should be used for quantitative real-time PCR, while RT-qPCR should be used for reverse transcription-qPCR, and genes used for normalization should be referred to as ...
Because of this, it is important to ensure that puppy diets contain higher amounts of energy than adult dog foods. However, over nutrition associated with feeding ad libitum results in accelerated skeletal growth and weight gain leading to osteopenia, especially in large breed dogs. [8]
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