Biosensor Parts:
How did we use Synthetic Biology?
The design of these parts was made thinking about the functionality of the protein in the sensor we proposed. We found that the receptor protein ESR1 was ideal to capture EDCs, so we created the Biosensor Parts to achieve the goal of expressing ESR1, purify it and immobilize it. We will briefly explore the functionality of each part, as well as key aspects of its design.
BBa_K3809010: ESR1 with Linker, His Tag and Signal Peptide
Figure 1.
Map of BBa_K3809010. Created with Snapgene.
This sequence codes for the Human Estrogen Receptor Alpha. It was modified in order to express the ligand binding domain and the DNA binding domain. The sequence also includes a 6X Histidine Tag (for its purification), a linker sequence of 4 glycines, 1 serine and 1 cistein (for its immobilization) and a signal peptide sequence (NPS4). For more information, click on the part name.
BBa_K3809011: mRFP with Linker, His Tag and Signal Peptide
Figure 2.
Map of BBa_K3809011. Created with Snapgene.
This sequence codes for protein reporter mRFP. It was modified in order to express the complete sequence of mRFP, with some additions. The sequence also includes a 6X Histidine Tag (for its purification), a linker sequence of 4 glycines, 1 serine and 1 cistein (for its immobilization) and a signal peptide sequence (NPS4). The purpose of expressing this protein is to analyze its folding and correct immobilization. For more information, click on the part name.
BBa_K3809012: Modified ESR1 protein under strong constitutive promoter and RBS
Figure 3.
Map of BBa_K3809012. Created with Snapgene.
This sequence is the result of the assembly of two parts: BBa_K3809010 and BBa_K081005. The first one is the coding sequence of the Human Estrogen Receptor ESR1. The second part is a strong constitutive promoter with an RBS with an efficiency of 0.6. We used this part for the expression of the modified ESR1 protein and its subsequent purification so that we could use it for the detection of Endocrine Disrupting Chemicals or EDCs. For more information, click on the part name.
BBa_K3809013: Modified mRFP protein under strong constitutive promoter and RBS
Figure 4.
This sequence is the result of the assembly of two parts: BBa_K3809011 and BBa_K081005. The first one is the coding sequence of a modified mRFP1. It was modified in order to express the reporter protein alongside other features we added to prove the correct folding of mRFP and ESR1. The second part is a strong constitutive promoter with an RBS with an efficiency of 0.6. We used this part for the expression of the modified mRFP protein so that we could use it to test the correct folding of the reporter, as well as to check if the immobilization in a Quartz Crystal Microbalance could be done with the characteristics we added. For more information, click on the part name.