Determination of Transgene Copy Number by Real-Time Quantitative PCR

Date
2009-01-01
Authors
Shepherd, Colin
Moran Lauter, Adrienne
Scott, Marvin
Scott, M. Paul
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Agronomy
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Agronomy
Abstract

Efficient methods to characterize transgenic plants are important to quickly understand the state of the transformant. Determining transgene copy number is an important step in transformant characterization and can differentiate between complex and simple transformation events. This knowledge can be extremely useful when determining what future experiments and uses the transgenic lines can be utilized for. The method described here uses real-time quantitative PCR to determine the transgene copy number present in the genome of the transformant. Specifically, this method measures the relative transgene copy number by comparing it with an endogenous gene with a known copy number. This method is a quick alternative to the Southern blot, a method that is commonly used to determine gene copy number, and is effective when screening large numbers of transformants.

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This is a chapter from Shepherd, Colin T., Adrienne N. Moran Lauter, and M. Paul Scott. "Determination of transgene copy number by real-time quantitative PCR." Transgenic Maize: Methods and Protocols (2009): 129-134, doi: 10.1007/978-1-59745-494-0_11.

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