荧光定量PCR技术使用者必读的文献－系统阐述荧光PCR技术扩增效率和使用范围的文献
丁香园论坛
1800
Validation of a quantitative method for real time PCR kinetics
Abstract
Real time RT-PCR is the most sensitive method for quantitation of gene expression levels. The accuracy can be dependent on the mathematical model on which the quantitative methods are based. The generally accepted mathematical model assumes that amplification efficiencies are equal at the exponential phase of the reactions for the same amplicon. However, no methods are available to test the assumptions regarding amplification efficiency before one starts the real time PCR quantitation. Here we further develop and test the validity of a new mathematical model which dynamically fits real time PCR data with good correlation (R2 ¼ 0:9995 0:002, n ¼ 50). The method is capable of measuring cycle-by-cycle PCR amplification efficiencies and demonstrates that these change dynamically. Validation of the method revealed the intrinsic relationship between the initial amount of gene transcript and kinetic parameters. A new quantitative method is proposed which represents a simple but accurate quantitative method. 2002 Elsevier Science (USA). All rights reserved.
Keywords: Real time PCR; Kinetics; Mathematical model; Quantitation of gene expression
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Abstract
Real time RT-PCR is the most sensitive method for quantitation of gene expression levels. The accuracy can be dependent on the mathematical model on which the quantitative methods are based. The generally accepted mathematical model assumes that amplification efficiencies are equal at the exponential phase of the reactions for the same amplicon. However, no methods are available to test the assumptions regarding amplification efficiency before one starts the real time PCR quantitation. Here we further develop and test the validity of a new mathematical model which dynamically fits real time PCR data with good correlation (R2 ¼ 0:9995 0:002, n ¼ 50). The method is capable of measuring cycle-by-cycle PCR amplification efficiencies and demonstrates that these change dynamically. Validation of the method revealed the intrinsic relationship between the initial amount of gene transcript and kinetic parameters. A new quantitative method is proposed which represents a simple but accurate quantitative method. 2002 Elsevier Science (USA). All rights reserved.
Keywords: Real time PCR; Kinetics; Mathematical model; Quantitation of gene expression
文献链接：http://www.transhold.net/services.htm
留个msn方便交流：transhold@transhold.net QQ技术讨论群：13893727
