Team:XMU-China/brainstorm

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MINUTES OF THE FIRST BRAINSTORM MEETING

held at Chemistry Building 104
at 7:00pm, Mar 10th ~11th , 2012
In attendance: Zhaoshou Wang, former members &current members of XMU iGEM
To discuss the feasibility of all the ideas raised by members

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Group 1 : regulating cell regeneration

Speaker: Muxin Yu, Ruosang Qiu, Yunxin Long
1.-Yu
Cell aging is related to telomerase. Telomerase may help supply a gap of the DNA synthesis mechanism. Research about cancer cell activation.
2. -Qiu Expands our project in 2011
a) Using AiiA to restrain AHL synthesis
b) CcdA&CcdB：CcdA >CcdB, function of CcdB is restrained；CcdA<CcdB CcdB inhibit DNA gyrase
3. -Long
The concept of Rehabilitation, Stem cell, iPS cell, Cell Reprogramming. But there would be much obstruction using stem cells or reprogamming cells to perform iGEM experiments.
Conclusion:
Considering our experiment condition, the second idea is most practical. To control cells density, we can use CcdA to restraint CcdB and then release the DNA gyrase astricted by CcdB. Compared with site-directed mutagenesis and sequentialermr-PronePCR, controlling cells density in such way would be more labour-saving.

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Group 2:Bioscreen

Speaker: Qingshu Wu, Yizhen Yan, Yuan He
1. –Wu
Literature summarize
a) Circuit with Boolean logic:simple circuitintricate circuit
b) About signal molecule
2. –Yan
a) Binary transfer encoder NAND gate digital tubes
b) Density display device: A/D converter encoder digital tubes
Special difficulties: constructing an operational amplifier
3. –He
a) How to decompose fluorescent proteins in order to recycle the displayer device? One of the idea is to use pigment to excitation fluorescence, then add some enzymes to decompose it.
b) Cells immobilization embedding cells
Conclusion:
Some intricate circuit may be too long to express, in this case, connecting expressive gene with promoter in series or in parallel can help. To construct recycling fluorescent device, fluorescent proteins should decompose themselves. How can we attain that? After studying a lot of literatures, members knew more about biocomputer, and about how to transfer signals on biologic units. Yan simply drew a logic circuit which turns input signals (1 or 0) into corresponding numbers (1-9). In that way, our plan of constructing a display device will be easier to achieve. To fix the fluorescent cells, they primarily decided to embed cells in special tubes. Yet how to switch the number show on the screen, namely how to decompose fluorescent proteins is the primary problem. Group 2 and group 7 have started finding a solution to this problem.

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Group 3: Constructing engineering bacteria to produce cellulase

Speaker: Chaoqun Hu, Xinzhu Tong
1. It's hard to make use of cellulose
2. Cellulose—endoglucanase, exoglucanase, β-1,4-glucosidase
3. The case of University of Edinburgh in 2007: Intracellular proteins are not able to excrete by themselves. How to solve this problem?
To do research about:
a) Ways of cells secreting proteins
b) How do original cells excrete cellulose?
c) In nature, how do living beings utilize cellulose ?
What's the advantages of this plan?
Conclusion:
After studying literatures, members of group 3 learn about ege gene and Bgat1 gene that published in2012, which can be used to produce cellulase. Generally in the process of producing cellulose, some enzymes must be added for lysis, and then release cellulase out of cells.
Group 3 members hoped to utilize the device controlling cell density which were constructed last year, and exchange ccdB to another gene which will let cells decompose themselves. Thus our engineering bacteria will not only produce cellulase with quorum sensing, and release it out of cells. Besides, group 3 are trying to construct a secreting routeway, hoping to produce cellulase without cell discruption.

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Group 4: photosynthetic bacterium(PSB) utilization

Speaker: Zhao Ma, Hong Sun, Youbin Mo
1. –Ma
a) photosynthetic bacteria
b) Mechanism of photosynthesis of purple bacterium
c) The case of California Institute of Technology in 2011
2. –Sun
a) Cyclic photophosphorylation
b) Nitrogen fixation
3. –Mo
a) Metabolism of acrylamide
b) Utilize of light energy: bacteriaamylose/proteinenergy
c) Finding usable bacteria
Conclusion:
Group 4 firstly introduced photosynthesis and its 3 major metabolic pathways. It's arduously to clone these pathways, so they are considering constructing some biobricks, which help to transform the organics that photosynthetic bacteria cannot utilize to another matter that can be utilize. Thus some organics can be decomposed by our engineering bacteria.

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Group 5: Utilizing engineering bacteria of quorum sensing (QS) system to increase Natamycin production

Speaker: Rong Fan, Qian Liang, Xinyi Yao
1. –Fan
a) Mechanism of bacteria quorum sensing (QS)
b) GBL(Gamma-butyrolactone) signaling molecules
c) About Natamycin
2. –Liang
a) Regulatory genes—pimM and pimR
b) High-efficiency production of Natamycin
Key points:
1) Is it viable for mass-producing?
2) Negative feedback& positive feedback
3) Association of quorum sensing and pimM expression
Conclusion:
Group 5 mainly searched information about increasing the production of Natamycin. But they didn't study about E coli's intracellular expression. Besides, they didn't mention of the advantages of utilizing quorum sensing (QS) system to produce metabolite. Therefore, in the following days, group 5 members should find answers to the two questions through studying literatures.

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Group 6: Secretive expression of recombinant Barnase by E.coli

Speaker: Que Huang, Zhibin Gu
1. barnase，barstar, RNase
2. Cancer cellsacidityseparationdeath
Normal cellsbasicity non separation
3. Cloning strategies of Barnase gene high fidelity PCR, why any error? 4. Secretive expression and Kil-Km Secretive box
Conclusion:
Barnase is a kind of RNase. It's toxic and it will cause cells death. Barstar is the depressor of Barnase. These two proteins interact and then neutralize the toxicity of Barnase. Group 6 members introduced barnase and barstar from various aspects, and told their applied prospect. Whereas they didn't learn about any problem in the process of producing Barnase, so we don't know what Synthetic biology can help to improve it. What's more, just to construct a biobrick to produce Barnase seems of not much value.
After discussion, we suggest group 6 members study about some difficulty in the productive process of Barnase, and try to communicate with group 5, finding our advantages of utilizing quorum sensing. Besides, Huang introduced some information of Kil-Km Secretive box, which can accelerate secreting proteins out of cells. So we hope them to exchange information with group 3 and help them to find secretive pathways.

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Group 7: Modulating the concentration of tricolor fluorescent proteins to show different colors

Speaker: Zhenan Zheng, Sifan Wang, Jianzhao Chi 1. –Zheng
Fluorescence resonance energy transfer (FRET)
a) Dual color FRET: Constructing fusion proteins are required
b) Multicolor FRET: To detect base mispairing
2. –Wang
Unstable fluorescent protein
To add peptide tail after GFP so as to be identified and decomposed by intracellular proteinase. The key lies in controlling and modulating the fluorescence intensity and the degradation velocity.
3. –Chi
Function of some Nano antibiotics: Modifying GFP's character.
Conclusion:
Study of group 7 focused on the choke point of this project: The degradation of fluorescent proteins, i.e. how to switch from one color to another. This Problem was also mentioned by group 2. How to switch from number A to number B when constructing a bioscreen? After studying literatures, group 7 put forward three probable proposals.
1) FRET mechanism
2) Unstable fluorescent protein
3) Utilizing Nano antibiotics to regulate fluorescent protein
After discussion, we thought the proposal two is relatively viable. For proposal one, mutual interference between yellow and blue lights is a hidden trouble. And the catabolic enzymes of Nano antibiotics mentioned in proposal three may be not easy to find.
So group 7 is suggested to study further about unstable fluorescent protein and peptide tail. But all of the three proposals are worth trying, and group 7's study results can act as reference of group 2's project—bioscreen.

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Group 8: Utilizing aptamers biobrick to construct biosensor

Speaker: Shuqin Hu, Mouzhe Xie, Zebo Hu
1. – Shuqin
a) Aptamers: widespread use in electrochemistry field, but it's application in biosome is still in initial stage.
b) Estrogen sensor
2. –Mouzhe
in vitroin vivo
What's the relationship between the biosensor and Synthetic biology? a) Constructing probe molecules. Whereas it may be difficult to construct, because single chains are not stable
b) Application in microbe. Introducing unstable single chains may lead to degradation.
3. –Zebo
Feasibility analysis
a) SELEX (systematic evolution of ligands by exponential enrichment)
b) Comparing aptamers with antibiotics
Better in stable, affinity; more extensive and more sensitive targets.
Conclusion:
Group 8 members told us the definition of aptamer and biosensor, and introduced the project of Pro. Yang Zhaoyong's group. Whereas they found that aptamer biosensors and Synthetic biology have little conjunction. At present, aptamer biosensors are usually selected in vitro, for example attaching on surface of an electrode.
If we want to do research of aptamer biosensors in vivo, there will be two ways:
1) To construct it in microbe, and let it transcribe single chains we need for making biosensor. But single chains are not stable. So this plan needs further research.
2) To select aptamers we need in vitro, then transform it in vivo, meanwhile its function of sensor should be maintained.
After discussion, we thought the two means are both of great difficulty. Group 8 can ask teacher Wang and Pro. Yang for help, seeking for the bonding point of aptamer biosensor and Synthetic biology or biobrick.