Team:LMU-Munich/Team/Collaborations

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Revision as of 10:55, 10 October 2014

Collaborations

Collaborations and the exchange of knowledge were important aspects during our iGEM participation. Therefore we had a intensive collaboration with the iGEM team Groningen (Netherlands) and laid a high priority on the exchange with the Rathenau Institute also located in the Netherlands. In addition we also took part in the Interlab Study of this year´s iGEM competition.

Interlab Study

Background:

The aim of the Interlab study was to evaluate three devices consisting of different Anderson promoters fused to GFP in two different backbones: pBS1C3 and pBS3K3. The devices were distributed by iGEM and the flourescence evaluation method was free of choice. For more details check: Interlab Study@iGEM

The devices:

  • Device one: BBa_I20260 consisting of J23101-B0032-E0040-B0015 in the backbone pBS3K3
  • Status: ready for evaluation
  • Device two: BBa_J23101 + BBa_E0240 in the backbones pBS1C3
  • Status: cloning before evaluation
  • Device three: BBa_J23115 + BBa_E0240 in the backbones pBS1C3
  • Status: cloning before evaluation

The cloning:

Devices two and three were built from their individual parts. By digesting the promotors (BBa_J23101and BBa_J23115) with SpeI-HF and PstI-HF restriction enzymes the backbone was cut open. The XbaI and PstI-HF digested GFP (BBa_E0240) was then ligated behind the promotors. After that both devices were transformed into DH5α, plated out on chloramphenicol [35mg/µl] LB-agar plates and tested via colony-PCR to identify positive clones. For clarification PCR-positive clones were sequenced: BBa_J23101 + BBa_E0240.txt
BBa_J23115 + BBa_E0240.txt

Flow through cytometry analysis:

For one measurement 50 µl of an overnight culture was first incubated together with 1 µl FM4-64 at 37°C to later on identify life/dead cells. After that 1 µl of each sample was diluted in 200 µl phosphate buffered saline solution and then evaluated by flow through cytometry. In order to generate meaningful results the described measurement was perfomed three times with technical replicas. For complete details please check our Interlab study submission: LMU-Munich Interlab form

Results:


LMU14 Interlab 1 091014.png GFP fluorescence LMU14 Interlab 2 091014.png





Discussion:





Collaboration with Team Groningen

Hi there!

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