Quantitative real-time PCR as a screening tool for estimating transgene copy number in WHISKERS™-derived transgenic maize

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Abstract

Quantitative real-time PCR (qRT-PCR) was adapted to estimate transgene copy number in transgenic maize callus and plants. WHISKERS™-derived transgenic callus lines and plants were generated using two different gene constructs. These transgenic materials represented a range of copy number. A 'standard curve' was established by mixing plasmid DNA with non-transgenic genomic maize DNA using a calculated ratio of target gene to host genome size. 'Estimated' copy number in the callus lines and plants using qRT-PCR was correlated with the 'actual' copy number based on Southern blot analysis. The results indicated that there was a significant correlation between the two methods with both gene constructs. Thus, qRT-PCR represents an efficient means of estimating copy number in transgenic maize.

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Song, P., Cai, C. Q., Skokut, M., Kosegi, B. D., & Petolino, J. F. (2002). Quantitative real-time PCR as a screening tool for estimating transgene copy number in WHISKERSTM-derived transgenic maize. Plant Cell Reports, 20(10), 948–954. https://doi.org/10.1007/s00299-001-0432-x

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