Team:Utrecht/Parts

Several biobricks from the kit were used during our project. Some of these were isolated from the kit by direct transformation of 2 μl of DNA resuspended in elution buffer NE. Besides the biobricks that were isolated from the kit, some constructs were ordered completely from IDT (Table 1). These Biobricks were used to create several new biobricks that were relevant to our project. BBa_K2736000 and BBa_K2736001, were used as BRET pair to measure the activity of our customized Tar receptors BBa_K2736003, BBa_K2736004, and BBa_K2736005. To test whether it was possible to switch ligand binding domains of the Tar receptor and two component receptors we created BBa_K2736002, and BBa_K2736006.

Used Biobricks

Table 2: Receptor EC50 values
Name	Biobrick Number	Size	Size	Obtained
RFP	BBa_K516032	828	Red fluorescence	isolated
Promoter	BBa_K608003	56	Strong promoter with medium ribosome binding site	isolated
eYFP	BBa_I15017	724	Yellow fluorescence	isolated
CheZ	BBa_K629003	645	"cheZ chemotaxis regulator protein phosphatase for CheY"	isolated
Copper promoter	98	Promoter responsive to phosphorylated CusR	ordered
Custom receptor	BBa_K2736006	1505 Fusion protein of CusS with the Tar ligand binding domain	ordered
1/5A::CheZ::Rluc	BBa_K2736002	1655CheZ fused to Rluc	ordered
CheY	390	Chemotaxis response regulator protein	ordered

Created Biobricks

Here comes Table 2