Team:Dundee/Parts/Ourbiobricks

Biobricks

We have submitted two BioBricks to the Registry of Standard Biological Parts that will hopefully be of use to future teams and projects.

1. BBa_K1012001 Protein Phosphatase 1

Human Protein Phosphatase 1 (PP1) is a protein from the family of serine/threonine phosphatases, we have used it as a microcystin binding protein however it regulates many processes in the body therefore it may be used in many other ways.

2. BBa_K1012005 ompC-GFP reporter construct.

This is an improved version of >BBa_R0083. BBa_R0083 comprises the ompC promoter, containing OmpR-binding sites. To improve this brick we added a strong Ribosome Binding Site (RBS; from BBa_B0034) followed by Green Fluorescent Protein (BBa_E0040). This was achieved by digesting BBa_R0083 with SpeI + PstI . The RBS from BBa_B0034 was excised with XbaI / PstI, and ligated into the BBa_R0083. The resultant plasmid was digested with SpeI + PstI , and was ligated with the GFP-encoding gene which had been excised from BBa_E0040 by digestion with XbaI / PstI. The resultant plasmid, Bba_ K1012005 responds to the osmotic activation of the EnvZ by producing green fluorescence. This part has been verified to work in this way.

In between the European and World Jamborees, we have successfully constructed the BioBricks we ran out of time when making for the European Jamboree.

The signal sequence of MalE

Signal allowing for protein transport via the general secretory pathway (Sec) in E. coli

BBa_K1012002 The TorA twin arginine transport signal sequence

DNA coding for the TorA (TMAO reductase) signal peptide. This part can be added upstream of a protein, targeting it for export by the twin arginine transport system (Tat) which transports folded protein. We used this part to export PP1 from the cytoplasm where it would fold to the periplasm of E. coli.

BBa_K1012004 The maltose binding protein (MalE) signal sequence

This is the DNA coding for the MalE signal peptide. Addition of this part at the 5’ end of a desired gene will target it for the general secretory pathway (Sec) which transports linear polypeptide. We used this part to target PP1 for the Sec machinery to transport it in to the periplasm of E. coli while we were investigating the best way to get PP1 there.