Team:HKUST-Rice/Phosphate Sensor

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Phosphate Sensor - P_phoA , P_phoBR

Potassium sensor

Nitrate sensor (p_dcuS)

Introduction

Phosphorus plays an essential role in plant growth. It associate with various growth factors for root development and seed production, etc. Deficiency in Phosphorus lead to stunted growth, yet symptoms are not obvious. Therefore, it is important to monitor the level of both the organic and inorganic phosphorus level in soil for healthy plant growth.

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Phosphate sensor Design

Escherichia coli detects inorganic phosphate (P(i)) from the environment by the PhoR/PhoB two-component system. As illustrated in Figure.1, P_phoA and P_phoBR is cross-regulated by PhoB and PhoR. The sensory histidine kinase PhoR can behaves as an activator or inactivator for PhoB depending on different states (inhibition state, activation state, deactivation state). When phosphate is limited, PhoR act as a phospho-donor for the autophosphorylation of PhoB. The phosphorylated PhoB will directly activate P_phoA and P_phoBR. In contrast, when there is phosphate, PhoR interferes with phosphorylation of PhoB which in turn inactivates the P_phoA and P_phoBR.

With the phosphate (Pho) regulon from E. coli,it can be utilized for detecting phosphate level. P_phoA and P_phoBR from E. coli strain DH10B were cloned and ligated with the GFP generator (pSB1C3-BBa_I13504) respectively. Under high phosphate concentrations, repression on the green fluorescence intensity is expected; while under low phosphate concentrations, expression on green fluorescence is expected.

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Results obtained

Characterization on the contructs (BBa_K1682013) using M9 minimal medium (without phosphate) were performed. Quantitative characterization on the promoters were done by measuring the fluorescence signal intensity using an EnVision multilabel reader.

The results were obtained by combining the 3 characterization results together.

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