Screening of transgenic plants by amplification of unknown genomic DNA flanking T-DNA

For the screening of transfer DNA (T-DNA) integration in transgenic plant m aterial, we developed a method based on specific amplification of genomic p lant DNA flanking T-DNA borders. This approach is possible because the leng th of the region flanking T-DNA extremity on a restriction fragment is spec ific to the integration locus. We have modified an adaptor ligation PCR tec hnique developed for amplification of unknown DNA flanking known sequence. The PCR patterns obtained were specific and reproducible for different plan ts from a given transgenic line. Furthermore, the number of PCR products ob tained could be considered a good estimation of the T-DNA copy number. When compared to Southern blot analysis, the PCR results give valuable compleme ntary information about the complexity of the T-DNA integration pattern and also about the integrity of the T-DNA borders. We describe the application s of this approach to populations of transgenic Arabidopsis thaliana plants .