High Frequency of Single-Copy T-DNA Transformants Produced After Floral Dip in CRE-Expressing Arabidopsis Plants

Transgenic plants that harbor a single copy of the introduced transgene are preferable to those with multiple transgene copies because multiple T-DNA copies correlate with expression variability and susceptibility to silencing. Especially after the commonly used floral-dip Agrobacterium -mediated transformation method, the frequency of single-copy transformants is low. The CRE/loxP recombinase-based strategy to resolve complex T-DNA loci has proven to be successful to efficiently obtain single-copy T-DNA transformants by directly transforming loxP -containing T-DNA vectors in CRE -expressing Arabidopsis thaliana plants. This chapter describes in detail how to transform three available loxP -containing T-DNA vectors into CRE -producing Arabidopsis C24 plants and subsequently how to analyze the transgenic plants for the T-DNA locus structure.