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External Cell Control Polymerase Chain Reaction: Replacing Internal Standards with an Unbiased Strategy for Quantitative Polymerase Chain Reaction Normalization by András Bors; Polett Ribiczey; Gabriella Köblös; Anna Brózik; Zsuzsanna Újfaludi; Mária Magócsi; András Váradi; Attila Tordai; Tünde Kovács; Tamás Arányi is a Biochemistry, Genetics and Molecular Biology article available to read on EtoBox.

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One of the most important strategic steps in the design of quantitative PCR experiments is the choice of an adequate internal standard gene. A reliable standard gene is expected to show unchanged expression under all experimental conditions. If the mRNA level of the standard gene is altered during the experiment, small changes of target mRNA levels can be especially difficult to detect. Indeed, several studies reported that the expression level of commonly used internal standards (e.g., glyceraldehyde-3-phosphate dehydrogenase; GAPDH 2 ) may change considerably in response to various factors [2]. Despite that, the expression of standard genes is rarely controlled, introducing thereby a risk of misinterpretation when analyzing real-time quantification experiments. Several methods to avoid quantification errors due to the use of internal standards of variable expression have been developed recently. Nevertheless none of them solved the problem conclusively 2,]. Here we describe the external cell control PCR (eccPCR) method, an approach developed primarily for cell culture experiments. This techique controls all steps of sample preparation and overcomes the incertitude of normalizatio

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Author

András Bors; Polett Ribiczey; Gabriella Köblös; Anna Brózik; Zsuzsanna Újfaludi; Mária Magócsi; András Váradi; Attila Tordai; Tünde Kovács; Tamás Arányi

Publisher

Elsevier Science; Elsevier ; Elsevier Inc.; Elsevier BV (ISSN 0003-2697)

Published

2008

Language

EN

Field

Biochemistry, Genetics and Molecular Biology (Life Sciences)