Team:DTU-Denmark/Notebook

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You can't spell funding without fun... (Annemi) - 25/06/10

I had a look at some funding and where we should go from here.

  • We should definitely apply for the COWI fund. This will take some time since they have a lot of requirements for applying. (see http://www.cowifonden.dk/Vejledning_for_ansoegere.asp)
  • The ‘Fondeliste’ has been updated according to the funds that Anastasiya and Anja found in the beginning of June.
  • A document called ‘Company applications’ can be found in the ‘company’ folder. Please write in that document if you send out a company application.
  • An application to the fund ALECTIA has been send.

Birthday and fun day at the lab? (Maya, Anja, and Patrick) - 24/06/10

So today was yet another day in the lab, only difference being it was my birthday (Patrick). :D
The serial dillutions we plated of our transformants didnt seem to produce a substantial amount of colonies after having plated them yesterday, so we plated a 10-1 and a 10-2 dilution to get a higher number of colonies.
We purified plasmids from strain SP44, SP45 and SP46 using the miniprep kit, and ran the purified plasmids on a gel to check if our plasmid purification was efficient and successful. There is now a miniprep protocol in the protocol folder.
We also made some -80 freezing cultures of strain SP44-46. To keep track of the future freezing cultures we have made a document called strain bank which we should all use when we make some -80 cultures. It's in the research group-> strain and plasmid folder.
The O/N of SP58 didn't show any growth, so a new O/N was started.

Progress (Maya, Anja, and Patrick) - 23/06/10

So we started in the lab yesterday after having gotten all the needed information from Flemming concerning the FP's (fluorescence proteins).
We started constructing some template plasmids with FP's and markers, and so far its been a fun experience thanks to Hassan.
As things are now we have decided to use the fluorescent proteins mCherry and YGFP as the reporters, though the final template plasmids will include the fast-degradable fluorescent proteins of mCherry and YGFP.
Today we started O/N cultures of the following strains: SP44, SP45, SP46 and SP58. SP44-46 contains plasmids with CFP, yGFP and Cherry, and SP58 contains a plasmid with the markers CAM and KAN.
We also made a transformation using electroporation. We transformed plasmid pSLD3 into E.coli strain . pSLD3 is to be used as a recipient plasmid of markers and FPs. After the transformation, we made a serial dilution of the transformed strain and plated these. We have uploaded a transformation protocol in the research group folder- protocol folder.

iGEM the last 2 weeks - 21/06/10

So the last 2 weeks have been quite hectic and busy, and if you're wondering why there hasn't been an update its because alot of things keep coming up while working on the design.
The in-silico design is moving along well, though when every new step comes along we run into issues, just like today we realized that the UV system just might not work after all, long story short - because the excitation wavelength will interfere with the excitation wavelength of the red-light receptor. Solution: good old lov-tap will most likely come into the picture now.
In terms of the reporter genes, we were lucky enough to find Flemming back at CSM today, and so Maya and Juliet managed to get alot of good info from him about flourescence proteins which made us wiser in that area, although we also realized they arnt as straight forward to work with as initially thought, so we need to put in more work in that area as well.
We are set to get into the lab tomorrow to possibly start construction of the template plasmids with the flourescence proteins, that is of course if we have the FP's we want to work with and whether those have the appropriate degradation time.
That's about all for now, stay tuned for more exciting news in the days to come.

Maya, Anja, Malthe and Patrick's log - 08/06/10

We have finished the step-wise construction of our system, and have begun on the in-silico model.
We have uploaded the initial work from the in-silico model in a newly created folder in the research group called step-wise construction and in-silico model.

Maya and Patrick's log - 07/06/10

So today we have been working on the step-wise construction of our system, taking into account how to test each part along the way. We learned a lot about recombineering and the factors entailed in the process and have come up with the initial version of how we are going to construct the system.
As a rule of thumb we also learnt the following combinations and bad-combinations when working with FP's:

  • RFP can be used together with either GFP or CFP; and YFP together with CFP.
  • The bad combinations which we should never ever do are the following: RFP with YFP; and YFP with GFP.
We have also created a new folder in the research group in dropbox, called "Synthetic biology" where we have uploaded 2 articles about the first constructed synthetic cell, which we think would be a good idea if everyone had a look at. :)
We have also uploaded Sebastien's presentations from last week, and they can be found in the folder "Presentations from Sebastien" in the research group folder as well.
So, from all the work today we are trying to make a scheme of phases / phase planning of the construction parts that need to be completed in a described order that will be uploaded possibly within this week.

Captain's log 040610

Maya, Thomas, Patrick, Lisa and Annemi have been working on a project description for Sebastien. The document 'project description' can be found in dropbox in the research group folder. From now on this document will be the only one describing the project, therefore you should edit in the document and not create a new one. We have decided that every time we are doing something iGEM related, that it should be posted in this document. Please write the date and your name at the beginning of your log. This will make it easier for all of us to keep track of what is going on in the group. We have tried to make a simple plan of what needs to be done before going to the lab:

  • Investigate the E. coli chromosome to find out where we can integrate the switch
  • Find the BioBricks we will be using (DNA sequences)
  • Install the software CLC main workbench (the program which will be useful for the design of the system)
  • Design the system: Order of genes, restriction sites, DNA sequences, order of integration of genes into the plasmids
  • Which plasmids (Sebastien)
  • Primer design
  • How to test that the genes have been inserted (antibiotic resistance)
  • Make a phase planning and talk to Peter (divide into smaller parts and and test these individually)

Maya will start creating an illustration for the system during the weekend. Thomas will delete all irrelevant budgets from the dropbox. He will also send the project description to Sebastien.

Lab Work and Notebook

On this page is described the experiments, procedures and protocols that we have used.
Further the Results from relevant experiments are presented. We have written succeses as well as failures to share our experience, knowledge and tips n' tricks we learned while working with our BBrick parts and the BB-standards and methods.

Protocols
For all the methods we have shaped the protocols to the standards and experiences of our lab. We have collected the protocols we used in a comprehensive list below where it is possible to read them in full length. They contain our procedure as well as references. Some time this might be weakly documented when given to us by communication with supervisors.

Generel experiences

General experiments and procedures that both groups have used.

Biobrick assemply standards

standard.
3A.

PCR

Repressor group

more specific characterizing experiments the biobrick

Work flow

XXXX maybe show pictures of our work plan, or write it simplified - the more pictures the better XXX

Experiment I

what method what results

Experiment II

what method what results

Terminator group

XXXX here we should write a short abstract XXXX

Work flow

XXXX maybe show pictures of our work plan, or write it simplified - the more pictures the better XXX

Experiment I

what method what results

Experiment II

what method what results

Protocols

List of the protocols we used, and links to the word documents We should have references in our protocols, when available.

References and resources

  • xxxxxxxxxxxx