18th, Feb

Place: Room 363, Life Science building

Instructor: Mr.Hong

Recordist: Wuyang Chen

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I. Discuss about our project:
1. SNS:
To complete this project is hardly possible. While it is likely to realize part of it, such as a system which is able to create and sense signals. But we need to confer our project some new meanings.
1) About making a signal disappear, according to Mr. Hong, we can add a rapid degradation label onto the signal.
2) We are afraid that the colony is too small to create an appropriate concentration gradient. According to Mr. Hong, it won’t be a big problem.

2. Solve the graph colouring problem using E.coli
1) In fact, we will only use E.coli to emit light in different colors, and all the algorithm for solving graph coloring problem is processed artificially.
2) We need to consult professors in Chemistry Department about constructing structures using ssDNA.
3) The scale of DNA strand is far too small than that of E.coli.
4) The fusion protein of Zn finger and the Ag43 would be very useful. We can think about how to make use of it.

3. Fatigue of specific stimulus
1) comK is a protein produced by bacillus subtilis and “competence” is a state of it. The possibility of turning into competence state is proportional to the amount of comK. We can substitute the protein involved in competence state for the protein we need.
2) The system can be used as a special lock.
3) How to make circuit one recover its function as quick as possible?
a) by means of protease?
b) by means of aptamer?

II. Set up groups to continue brainstorm

III. Make plans for the whole term.

25th, Feb

Place: Room 363, Life Science building

Instructor: Mr.Hong

Recordist: Xingwen Chen

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Continue brainstorming.
I.Three ideas we have put forward before:
1.Bacterio-SNS (Social Networking Services )
problem1.The signal molecules may exist for too long.
solution：Add a degradation tag to the signal protein for the cell to degrade it quickly.
problem2.How to construct a memory system?
There are several ways but which is the most suitable? problem3.Does CDK exist in bacteria?
We can't get it until we find a paper about that.
problem4.We should consider what kinds of result we will achieve.

2.Solve graph coloring problem with DNA and visualize the result with E.coli.
problem1.How to construct the complex DNA structure?
We can ask a professor who are studying this.
problem2.Bacteria are much bigger than DNA,thus the visualization may be impossible.

3.E.coli mimic specific stimulation fatigue
problem1.The idea will actually be quite boring if our purpose is only to mimic.
Think of something new which can apply the system.
problem2.The signal molecules may exist for too long and signals may influence each other.
Change the signal and consider some physical signals such as temperature and light.


II.New Ideas
Zhou Shan says:
1.Bacteriaphage therapy

2.Synthetic biology moving into the clinic

3.improve bacteria tolerance：
a)efflux pump
b)heat shock proteins
c)membrane modification
d)general stress respond
These are mostly achieved by scientists.It is wrong to repeat others unless we can come up with something new.

4.Some thoughts about nucleic acid aptamer
a) Aptamer itself can be substrate.
b) Two aptamers cna be combined on a single bead.


III.Professor Liu suggest we could do something on the basis of our work of iGEM 2011.
Luo Siwei, a team member of iGEM2011,says that if we must do as this,the only thing we can do is to make it more specific,which seems a little boring.And he indicates that the work last year is too complicated and lacking in aesthetics.


IV.About aptamer
We have the technology and we can start the SELEX now.But we have to work out a good application of it.


V.Tasks of the next two weeks
Direction 1.Come up with a new idea on the basis of the work last year.
a)What can we improve?
b)Think about uncompleted work.
c)Try to apply the signal and response system we came up with in the SNS subject.
d)Try to apply the circuits in the specific stimulation fatigue subject.
e)Read more papers,view previous works and get some inspirations.

Direction 2.Bacterio-SNS:
a) Learn more about toggle switch.
b) Find out if there is something similar to CDK complex in E.coli or B.subtilis.
c) Think about what effect shall we get finally.
Direction 3:E.coli mimic specific stimulation fatigue:
Think of an application of it.

July 10-July 16

7.10

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ligate aptamer-CheZ with PSB1C3 plasmids

extract plasmids containing LuxPR and terminator result:the concentractions of plasmids LuxPR and terminator are both 600ng/ul

7.11

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transform PSB1C3 containing aptamer-CheZ into bacteria and cultivate. 7.12

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carry out Double digestion of toggle switch and pSB1c3 and ligate them together.

pick up single colony from bacteria cultivated yesterday.

result: Both the location and the brightness of the electrophoretic bands correspond with our expectation.

7.14

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transform the part rbs-ci-ter sent from PKU again into bacteria. check the colors of the colonies containing toggle switch. result: the ratio of red vs green is 8:25, verifying the function of toggle switch.

7.15

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pick up the single colony from the plate yesterday and ten hours later extract the plasmids containing Ci-terminator.

7.16

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conduct PCR with the plasmids yesterday to reproduce rbs-ci-Term. result:failure

July 17-July 23

7.18-7.21 ---- ligate the standard part Terminator to the end of toggle switch result: we conduct a enzyme digestion with EcoR1 and Pst1 to the final ligation product and the electrophoresis result shows that the ligation is successful.

July 24-July 30

7.22-7.29 ---- perform the following experiments: PCR of rbs-ci-ter, enzyme digestion(E,S),ligation with LuxPR(standard part from IGEM registry digested by E,X),tranformation,plasmids extraction. result: the concentration of plasmids with ligation gene is as high as ...

July 31-Aug 6

7.30-8.3 ---- PCR of LuxPR-rbs-ci-ter, enzyme digestion(E,S),ligation with LuxPR(standard part from IGEM registry digested by E,X),tranformation,plasmids extraction.

Aug 7-Aug 13

Aug 14-Aug 20