Team:NTNU-Trondheim/Notebook/October

From 2013.igem.org

Trondheim iGEM 2013

October

Wednesday 02.10.2013

SDS-PAGE of bacterial samples with the tat_GFP_RFP (ER1) and tat_ProteinG construct

In order to determine if Protein G is produced in the S3-2B sample and wheater a GFP-RFP dimer is produced in the ER1 sample, a SDS-PAGE was performed. Liquid cell culture with ER1 (tat_GFP_RFP construct), S3-2B (tat_ProteinG construct) and wildtype ER2566 ''E.coli'' cells was centrifuged and the pallet was mixed with SDS loading buffer. This mix was then heated at 95 °C for 15 minutes. The SDS-PAGE results can be viewed in the figure below

Figure 1: SDS-PAGE of bacterial samples. From the left: wild type ER2566, tat_GFP_RFP and tat_ProteinG

There is evidently produced Protein G in the S3-2B sample as there is an additional band at the expected size of about 60 kDa. There is also an additional band at the ER1 sample of about 28-30 kDa. This half the expected size of about 60 kDa. This indicates that only RFP is produced in the bacterias.

SDS-PAGE of bacterial samples with the tat_GFP_RFP (ER1) and tat_ProteinG construct

A new test on the Pm/XylS promoter was performed with the same conditions as earlier.

Wednesday 02 - Friday 04.10.2013

As the first attempt to send in the tat_GFP_RFP construct to iGEM Headquarters failed, we redid the cloning as we did 26-28.09.2013 and sent in a new construct.