Plant-derived insulator-like sequences for control of transgene expression

Stable and consistent transgene expression is necessary to advance plant biotechnology. Stable expression can be achieved by incorporating enhancer-blocking insulators, which are cis-regulatory elements that reduce enhancer interference in gene expression, into transgene constructs. Sufficient insulators for plant use are not available, and their discovery has remained elusive. In this work, we computationally mined the compact genome of Utricularia gibba for insulator sequences and identified short (<1 kb) sequences with potential insulator activity. Based on in vivo tests, three of these effectively mitigate the non-specific transgene expression caused by the Cauliflower Mosaic Virus 35S promoter. Our data indicate that these sequences are true enhancer-blocking insulators that can be used to reduce variable transgene expression in multigene constructs used for plant transformation.