Absolute Quantitative Real-Time PCR Assay for Determining Transgene Copy Number in Transgenic Mice
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R346

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    Abstract:

    Objective It is first and foremost to genotype and determine the transgene copy number for making a transgenic model.Southern blot analysis,the traditional means,is time consuming and laborious.The aim of this study was to built up a novel method to determine transgene copy number by using the absolute quantitative real-time PCR.Methods Taken TG-CARK founder mice as examples,the transgene copy numbers in transgenic mice were determined by quantitative real-time PCR and Southern blot.The conserved mouse housekeeping gene,Fabpi,served as an internal control to calculate transgene copy number.Results The copy numbers of transgenic founders estimated by real-time quantitative PCR were highly correlated with actual copy number based on Southern blot assay.The three TG-CARK founders had the transgene copies of 1,7,45 respectively.Conclusion Real-time PCR is a more efficient and convenient method for estimating copy number in transgenic mice than Southern blot.

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  • Received:
  • Revised:January 19,2007
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