A Suite of Constitutive Promoters for Tuning Gene Expression in Plants

The need for convenient tools to
express transgenes over a large
dynamic range is pervasive throughout plant synthetic biology; however,
current efforts are largely limited by the heavy reliance on a small
set of strong promoters, precluding more nuanced and refined engineering
endeavors in planta. To address this technical gap,
we characterize a suite of constitutive promoters that span a wide
range of transcriptional levels and develop a GoldenGate-based plasmid
toolkit named PCONS, optimized for versatile cloning and rapid testing
of transgene expression at varying strengths. We demonstrate how easy
access to a stepwise gradient of expression levels can be used for
optimizing synthetic transcriptional systems and the production of
small molecules in planta. We also systematically
investigate the potential of using PCONS as an internal standard in
plant biology experimental design, establishing the best practices
for signal normalization in experiments. Although our library has
primarily been developed for optimizing expression in N. benthamiana, we demonstrate the translatability of our promoters across distantly
related species using a multiplexed reporter assay with barcoded transcripts.
Our findings showcase the advantages of the PCONS library as an invaluable
toolkit for plant synthetic biology.