Background: FLP recombinase mediated integration into a pre-integrated FRT site is routinely used to generate highly reproducible stable transgenic cell lines. In this study, we broaden the system of site specific integration by introducing ΦC31 integrase mediated integration into attP sites.

Results: We generated a HEK293 host cell line with a single copy FRT as well as an attP site allowing site specific integration of two distinct transgenes. To achieve conditional control, we used the tetracycline and Shld1 inducible systems. By introducing fluorescent reporters we show that integration and induction of two transgenes are completely independent. We applied this new technique to investigate the effect of HNF4α on proliferation of HEK293 cells by introducing HNF4α into each integration site. We obtained in two independent cell lines highly reproducible results that prove the usefulness of this novel HEK-attP/FRT cell line.

Conclusions: In this study we have established and applied a HEK-attP/FRT cell line that allows site specific integration of two conditional transgenes using the FLP recombinase as well as the ΦC31 integrase.