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| | * Spent morning discussing ethics. | | * Spent morning discussing ethics. |
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| | * Lab Cleanup in the afternoon | | * Lab Cleanup in the afternoon |
| | * Worked on our ethics rough draft. I read over some ethics reports form last year (Heidelberg and TUDelft) to get some ideas for format/ structure. I re-wrote the introduction that we used for our last ethics assignment for AIF to have a better explanation of synthetic biology and our project in general. Fahd and I divided up sections to look at and add to this weekend (Economics and Legal for me and Social and environmental for Fahd) and then we will send it to Stefan to tie things together and make it more unified. We made a list of few papers that will be helpful in doing this. I also typed up some of my lab notebook to be used on the Wiki. | | * Worked on our ethics rough draft. I read over some ethics reports form last year (Heidelberg and TUDelft) to get some ideas for format/ structure. I re-wrote the introduction that we used for our last ethics assignment for AIF to have a better explanation of synthetic biology and our project in general. Fahd and I divided up sections to look at and add to this weekend (Economics and Legal for me and Social and environmental for Fahd) and then we will send it to Stefan to tie things together and make it more unified. We made a list of few papers that will be helpful in doing this. I also typed up some of my lab notebook to be used on the Wiki. |
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| - | 2) Called Julie Edwards at Boheringer Ingleheim Canada. She ws away from her office so i left her a voicemail. Will call her on Monday | + | 2) Called Julie Edwards at Boehringer Ingleheim Canada. She ws away from her office so i left her a voicemail. Will call her on Monday |
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| | - We have defined cell life cycle for the simulated cells and now they divide after specific amount of time. | | - We have defined cell life cycle for the simulated cells and now they divide after specific amount of time. |
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| - | - We can extract different kind of results such as concentration graphs, and matrixes, and rule charts (I will attach a figure of available results). | + | - We can extract different kind of results such as concentration graphs, and matrices, and rule charts (I will attach a figure of available results). |
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| | These components constitute the main framework of the system. Now we need a tutorial that helps biologists to understand this model. As a result, we have designed an animated tutorial which teaches the biological details of the system as well as the notations used to implement this system in Mathematica. One could look at these animations and realize how this model is related to the actual biological cascades of our circuits. | | These components constitute the main framework of the system. Now we need a tutorial that helps biologists to understand this model. As a result, we have designed an animated tutorial which teaches the biological details of the system as well as the notations used to implement this system in Mathematica. One could look at these animations and realize how this model is related to the actual biological cascades of our circuits. |
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| - | This figure demonstrates a view of this animation. This tutorials constitute one of the educational aspects of this project. | + | This figure demonstrates a view of this animation. |
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| + | The biological system goes through two main cascades. Each cascade has its own interactions and agents involved. A nice interface has provided for users to watch the animation of the cascades separately in respect to the interaction occurring. The following figure shows this interface: |
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| | Purpose: to move LuxPQ (insert) into the AC plasmid (vector) so that we can subsequently move B-R-OU-B into this plasmid. A restriction digest was set up by cutting the insert and vector with XbaI and PstI. After this restriction digest, phosphotase and ligation treatment was performed and the plasmid was subsequently transformed into TOP10 competent cells and plated overnight on LB+Chlor. | | Purpose: to move LuxPQ (insert) into the AC plasmid (vector) so that we can subsequently move B-R-OU-B into this plasmid. A restriction digest was set up by cutting the insert and vector with XbaI and PstI. After this restriction digest, phosphotase and ligation treatment was performed and the plasmid was subsequently transformed into TOP10 competent cells and plated overnight on LB+Chlor. |
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| - | In order to construct the PQ-B-R-OU-B circuit, another approach was used in which the LuxPQ in AK is inserted into B-R-OU-B in AC. This was set up by cutting the insert with EcoRI and SpeI and by cutting the recipient with EcoRI and XbaI. Again, after this restriction digest, phosphotase and ligation treatment was performed and the plasmid was subsequently transformed into TOP10 competent cells and plated overnight on LB+Chlor. | + | In order to construct the PQ-B-R-OU-B circuit, another approach was used in which the LuxPQ in AK is inserted into B-R-OU-B in AC. This was set up by cutting the insert with EcoRI and SpeI and by cutting the recipient with EcoRI and XbaI. Again, after this restriction digest, phosphatase and ligation treatment was performed and the plasmid was subsequently transformed into TOP10 competent cells and plated overnight on LB+Chlor. |
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| | The note cards for the ligation activity for molecular cloning have been created, consisting of parts from the registry: | | The note cards for the ligation activity for molecular cloning have been created, consisting of parts from the registry: |
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| | * J13002 – promoter + rbs | | * J13002 – promoter + rbs |
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| | After today’s meeting, I have sent Pqrr4+I13500 for sequencing with BBK CP F/R sequencing primers to verify if it is actually in the plasmid. After the verification, I will make it competent again so that Emily can test her mutants. It has been sent before 1pm, so I am hoping to get it by Monday. | | After today’s meeting, I have sent Pqrr4+I13500 for sequencing with BBK CP F/R sequencing primers to verify if it is actually in the plasmid. After the verification, I will make it competent again so that Emily can test her mutants. It has been sent before 1pm, so I am hoping to get it by Monday. |
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| | After lunch, we had a huge clean up time, as our lab was getting dirtier every day. | | After lunch, we had a huge clean up time, as our lab was getting dirtier every day. |
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| | We had a mini-meeting to list out all the commands and controls we thought users needed to know in the labs and the biobrick simulator so that Stefan could design some activities around teaching those. We also began to layout how we wanted the path in Synthetic Kingdom to work. Today I spent 5 minutes staring at Katie’s DNA Polymerase, which is SO CUTE. <3 | | We had a mini-meeting to list out all the commands and controls we thought users needed to know in the labs and the biobrick simulator so that Stefan could design some activities around teaching those. We also began to layout how we wanted the path in Synthetic Kingdom to work. Today I spent 5 minutes staring at Katie’s DNA Polymerase, which is SO CUTE. <3 |
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| | (shadowing Jeremy) I made a 0.7% agarose gel for restriction digest of Colony 6 and colony 7. (we took C6 +7 from a plasmid PCR that was done earlier by Jeremy to verify the presence of LuxPQ-B-R-OU-B in AC plasmid. C6 +7 looked the most accurate after Jeremy ran it on a gel so we used it for the restriction digest today. ) | | (shadowing Jeremy) I made a 0.7% agarose gel for restriction digest of Colony 6 and colony 7. (we took C6 +7 from a plasmid PCR that was done earlier by Jeremy to verify the presence of LuxPQ-B-R-OU-B in AC plasmid. C6 +7 looked the most accurate after Jeremy ran it on a gel so we used it for the restriction digest today. ) |
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| - | The purpose of this restriction digest was to verify the size of the PQ-B-R-OU-B. He let me do almost everything on my own while he was there. After we ran it on a gel, wells 2 and 4 (cut) was blank but the wells 3 and 5 (uncut) was clearly visible. We assumed there must’ve been a problem with the RD perhaps so we redid everything again. I made the gel again, etc. Yesterday, Jeremy did a RD of luxPQ and ccdb and left it overnight. Today we did a plasmid switch to transfer PQ from AK to AC. We anarctic phosphatsed it and ligated it, then we did a transformation to transfer PQ plasmid into top 10 cells. Finally we plated it on Chlroramphenical plates and left it overnight. | + | The purpose of this restriction digest was to verify the size of the PQ-B-R-OU-B. He let me do almost everything on my own while he was there. After we ran it on a gel, wells 2 and 4 (cut) was blank but the wells 3 and 5 (uncut) was clearly visible. We assumed there must’ve been a problem with the RD perhaps so we redid everything again. I made the gel again, etc. Yesterday, Jeremy did a RD of luxPQ and ccdb and left it overnight. Today we did a plasmid switch to transfer PQ from AK to AC. We antartic phosphatased it and ligated it, then we did a transformation to transfer PQ plasmid into top 10 cells. Finally we plated it on Chlroramphenical plates and left it overnight. |
| | The rest of the time was spent in the ethics meeting this morning and major lab clean up in the afternoon. | | The rest of the time was spent in the ethics meeting this morning and major lab clean up in the afternoon. |
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CAROL
Team Meeting and Lab Clean Up
- Spent morning discussing ethics.
- Lab clean up
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EMILY
Team Meeting, Lab Cleanup and Ethics
- Lab Meeting: Discussed Ethics Conference and paper
- Lab Cleanup in the afternoon
- Worked on our ethics rough draft. I read over some ethics reports form last year (Heidelberg and TUDelft) to get some ideas for format/ structure. I re-wrote the introduction that we used for our last ethics assignment for AIF to have a better explanation of synthetic biology and our project in general. Fahd and I divided up sections to look at and add to this weekend (Economics and Legal for me and Social and environmental for Fahd) and then we will send it to Stefan to tie things together and make it more unified. We made a list of few papers that will be helpful in doing this. I also typed up some of my lab notebook to be used on the Wiki.
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FAHD
Marketing and Ethics for July 21st 2009
Today, I concentrated my energies at marketing our iGEM project and looking at the ethical aspect of our project. Here is what I did today:
1) I called Edward Short at Charles River Labs. He is the HR director. Even though they said no, I am still on his case for contacts and sone money. I left a voicemail for him. Will call him on monday
2) Called Julie Edwards at Boehringer Ingleheim Canada. She ws away from her office so i left her a voicemail. Will call her on Monday
3) I called Mike Barr at Critical Outcome technologies. He is the Community Relations officer. He said that to call him on Wednesday July 29th.
4) I called Heather Bourdeu at life technologies. She is the community relations officer. Left her a voicemail and i will call her on monday.
5) I called Andrea Greene at EMD Bio-Chemicals. She is the Marketing/communications officer. Left her a voicemail and I will call her on monday.
6) I called Nic Milligan at Teck Coal Ltd. He is the community relations officer. Left him a voicemail and I will call her on monday.
7) I called Cindy Bizon at ALPAC. She is the community relations officer. Left her a voicemail and I will call her on monday.
8) I called Leon Zupan at Enbridge Pipelines Inc. He is the Director of Engineering and Development. Left him a voicemail and I will call him on monday.
9) I called Ron Hall at Focus Corporation. He is the Branch Manager. He wants our sponsorship packgae so I e-mailed him one. He is also an Alumini.
10) I called Dr Adolfo Cotter at Neuroimage Inc. for Contacts and Advice. Since his company is new, he will not be able to help us but he told nme to keep him in mind for next year and keep him updated. His advice was nothing new and was familiar to Derek Gratz's.
11) Helped clean the Lab.
12) Attended the meeting in the morning
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Review:
So, after almost 3 months working on this model, we have the following components:
- We have implemented the notations of rules, so that we can define various interaction rules for our system.
- We have functions that take in an interaction rule and apply it to the system.
- Gillespie's algorithm is implemented. As a quick review, this algorithm provides randomness for biological systems and also determines random time for each interaction.
- We have defined cell life cycle for the simulated cells and now they divide after specific amount of time.
- We can extract different kind of results such as concentration graphs, and matrices, and rule charts (I will attach a figure of available results).
These components constitute the main framework of the system. Now we need a tutorial that helps biologists to understand this model. As a result, we have designed an animated tutorial which teaches the biological details of the system as well as the notations used to implement this system in Mathematica. One could look at these animations and realize how this model is related to the actual biological cascades of our circuits.
This figure demonstrates a view of this animation.
The biological system goes through two main cascades. Each cascade has its own interactions and agents involved. A nice interface has provided for users to watch the animation of the cascades separately in respect to the interaction occurring. The following figure shows this interface:
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JEREMY
Plasmid switch moving LuxPQ from the AK plasmid to the AC plasmid
Purpose: to move LuxPQ (insert) into the AC plasmid (vector) so that we can subsequently move B-R-OU-B into this plasmid. A restriction digest was set up by cutting the insert and vector with XbaI and PstI. After this restriction digest, phosphotase and ligation treatment was performed and the plasmid was subsequently transformed into TOP10 competent cells and plated overnight on LB+Chlor.
In order to construct the PQ-B-R-OU-B circuit, another approach was used in which the LuxPQ in AK is inserted into B-R-OU-B in AC. This was set up by cutting the insert with EcoRI and SpeI and by cutting the recipient with EcoRI and XbaI. Again, after this restriction digest, phosphatase and ligation treatment was performed and the plasmid was subsequently transformed into TOP10 competent cells and plated overnight on LB+Chlor.
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KATIE
Notecards
The note cards for the ligation activity for molecular cloning have been created, consisting of parts from the registry:
- J13002 – promoter + rbs
- B0015 - terminator
I also made up a note card for a gene that was selected for one of the missions in the lab (AFP Gene) as well as note cards to give for each step of construction. Now I am in the process of renaming everything for restriction digest, phosphatase treatment and construction to work using these note cards and creating the substrings within the master note cards that I will need to compare with the note cards dropped into each object used for construction.
I also created a DNA polymerase for the DNA replication display that will move in a straight line and nucleotides will appear behind it as it moves. I also made a backbone for the strand that I was trying to create, but the backbone attached to each individual nucleotide does not line up perfectly together yet even though they are the exact same size so I will have to go back to the polymerase script to change the position that the nucleotides are rezzed at. I am aware that nucleotides are not produced by the DNA polymerase, but it is the only way to get the nucleotides to the position I want easily and it is hard to tell that they appear out of the DNA polymerase anyway. I am working on setting the position of the polymerase to a section on the leading strand of DNA that I also finished constructing today. I have also made helicase and DNA ligase, which presently do not do anything, but I will start working on the other enzyme’s functionality once I get the DNA polymerase running smoothly.
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KEVIN
1. Sequencing
After today’s meeting, I have sent Pqrr4+I13500 for sequencing with BBK CP F/R sequencing primers to verify if it is actually in the plasmid. After the verification, I will make it competent again so that Emily can test her mutants. It has been sent before 1pm, so I am hoping to get it by Monday.
2. Lab clean up
After lunch, we had a huge clean up time, as our lab was getting dirtier every day.
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MANDY
Fixing DNA Extraction and Further Planning in Second Life
Today, tested out the DNA extraction activity I did last week. It worked up until the last few steps, where I think I have my parts miscommunicating with each other and the centrifuge near the end, so I am in the process of working backwards to check all my scripts and change this. After this, I can make start on biobrick PCR, using the PCR that Katie has already scripted. This will allow us to work on the ‘restriction digest and ligation’ of the circuit that is required for the third lab mission. Once this is finished, we will evaluate if we have time to do cell culture for the first lab mission. Either way, the first mission has bacterial transformation which has been scripted in the form of a 10 question quiz by Katie, which is easier to understand than the equipment used in the more difficult levels, allowing the user to get used to dialogue box controls.
Once the lab missions are finished, we will clone the entire thing into the green island and make everything green. After the labs are complete, we will begin working on the base of Patrick’s biobrick simulator. Katie and I have a few notecards already written up on the basics, we just need to organize it in an easy to understand format with something creative that is more interesting than just reading notecards. Katie is working on having a moving ‘diagrams’ eg of DNA polymerase, etc. I am going to figure out how to embed streaming video in Second Life (maybe show a moving visualization of transcription).
We had a mini-meeting to list out all the commands and controls we thought users needed to know in the labs and the biobrick simulator so that Stefan could design some activities around teaching those. We also began to layout how we wanted the path in Synthetic Kingdom to work. Today I spent 5 minutes staring at Katie’s DNA Polymerase, which is SO CUTE. <3
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PATRICK
Working While Dodging Meetings
Retrospective Notebook: This entry was not written on this day, but derived later from working notes I made that day.
Unlike the title, most of today was spent in meetings, so little work on my corner of the project got accomplished as a result. But now I understand everyone else's work a little better!
Moving objects around in three dimensions is neat, but it can be a bit of a pain in second life. I worked on changing the way the objects bind and unbind so that they mostly stay in the same plane, which makes them much easier to deal with.
I also started planning what elements I would show when my turn to do a [http://igemcalgary.blogspot.com/2009/07/patrick-and-biobrick-simulator.html video blog] came around.
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PRIMA
Marketing & RD of PQ-B-R-OU-B
Today:
In Marketing:
I called Courtyard Marriot Hotel in Boston. They offered 279/night with 2 queen beds per room. I wanted to know if they offer any quotes for students but apparently Marriot hotels, in general, don’t offer any sort of quotes/discounts.
I mailed the sponsorship pkg to GSK (GlaxoSmithKline) just as the Charitable Donations lady asked me to. Although she didn’t think iGEM’s goal this year matches the company’s objectives but she’s still willing to have a look at it.
I didn’t call any companies today because I’ve been continuously calling them for the past few weeks. I’ll do it Monday though.
Company 1’s donation committee reviewed our pkg and said that since that company does not have a presence in Calgary and they only support communities where they have operations so they declined our proposal. I sent her a thank you letter and asked for contacts.
In lab:
(shadowing Jeremy) I made a 0.7% agarose gel for restriction digest of Colony 6 and colony 7. (we took C6 +7 from a plasmid PCR that was done earlier by Jeremy to verify the presence of LuxPQ-B-R-OU-B in AC plasmid. C6 +7 looked the most accurate after Jeremy ran it on a gel so we used it for the restriction digest today. )
The purpose of this restriction digest was to verify the size of the PQ-B-R-OU-B. He let me do almost everything on my own while he was there. After we ran it on a gel, wells 2 and 4 (cut) was blank but the wells 3 and 5 (uncut) was clearly visible. We assumed there must’ve been a problem with the RD perhaps so we redid everything again. I made the gel again, etc. Yesterday, Jeremy did a RD of luxPQ and ccdb and left it overnight. Today we did a plasmid switch to transfer PQ from AK to AC. We antartic phosphatased it and ligated it, then we did a transformation to transfer PQ plasmid into top 10 cells. Finally we plated it on Chlroramphenical plates and left it overnight.
The rest of the time was spent in the ethics meeting this morning and major lab clean up in the afternoon.
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STEFAN
Decisions, Decisions
Today was mostly meeting...
BUT our team decided on a bunch of Second Life stuff afterward. The
most important part was deciding what to teach people when they first
come to our island. In particular: flying, teleporting, opening your
own and other object inventories, and manipulating objects. Some of
these would be taught on the starting island, and others will be
taught in the Synthetic Kingdom (for practice). For example, you click
on a jellyfish to receive GFP and then open the inventory of a
bacteria, put it in, and then the bacteria glows. Speaking of that
wacky place, we brainstormed some rough ways to structure it. I think
it is still a good idea to have all the bacteria floating around to
give it a sense of awe, however, it could benefit from a path. This
would make it very similar to a zoo exhibition.
The path would preferably be lighted for maximum awesomeness
and each "station" would hold a different engineered bacteria. There
would also be signs pointing the way and a little notecard giver that
would give you more information.
This weekend will spent on the paper. Emily and Fahd were working on
it today and I'll complete it on Sunday.
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VICKI
Bringing credibility to my introduction
I found 4 papers that would be really useful: 2 that describe the AI-2 signalling pathway (Chen et al, 2008; Bassler et al, 1994); 1 that describes the AHL signalling pathway in depth (Cao et al, 1989); and some justification for why we’d bring in AI-2 in addition to AHL in that AI-2 is much more prevalent in nature, which should make it more versatile in its potential applications (Rasmussen, 2006).
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