Bioimaging of phosphate uptake and homeostasis in roots

We will use a protein-based nanosensor for phosphate to follow phosphate uptake in vivo with a confocal microscope  
 
A phosphate nanosensor consists of a substrate-binding protein linked to two fluorescent reporter proteins. Substrate binding changes the conformation of the nanosensor and the efficiency of fluorescence resonance energy transfer (FRET) between the reporter proteins. 

 

A Pi nanosensor has been engineered by fusing a predicted bacterial Pi-binding protein (piBP) from Synechococcus to eCFP and eYFP as FRET partners (Gu et al., 2006) and constitutively expressed Arabidopsis.

 

In this project we will test substrate specificity and root response on externally applied phosphate. e trying to measure Pi changes over a wide range (nanomolar to millimolar) with this genetically encoded Pi FRET nanosensor expressed in Arabidopsis.