PRELIMINARY MOLECULAR EVIDENCE THAT QUANTITATIVE POLYMERASE CHAIN REACTION DETECTS CHIMERAS IN TRANSGENIC PLANTS

Plants transformed via Agrobacterium tumefaciens are prone to regenerate a chimera, or mixture, of transformed and non-transformed cells.
In this study, we developed a method to identify putative chimeras in transgenic tobacco and apricot plants using quantitative real-time polymerase chain reaction (qPCR). The qPCR assay is based on the assumption that the amount of nptII DNA relative to an internal control (actin) DNA should be constant for solid, non-chimerical transgenics but will vary among tissue samples and over time for a chimerical clone. qPCR was used to estimate the amount of nptII DNA and actin control DNA that was used to normalize the amount of target DNA in each reaction.
This method was able to detect putative chimeras when multiple tissue samples were evaluated from the same line.
The method was successfully used to monitor the dissociation of chimeras in tobacco plants and apricot callus.
Although Southern blot analysis is widely used to confirm transgene integration, we demonstrated here that it cannot detect T-DNA insertion chimeras.

I International Symposium on Biotechnology of Fruit Species: BIOTECHFRUIT2008

M. Faize, L. Faize, L. Burgos

Chimera, qPCR, nptII, actin

https://doi.org/10.17660/ActaHortic.2009.839.46