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| | * Final transformation did not result in any colonies. The team has decided to use GFP as a reporter circuit instead since this transformation is not cooperating with us. | | * Final transformation did not result in any colonies. The team has decided to use GFP as a reporter circuit instead since this transformation is not cooperating with us. |
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| | Gauntlet replied back : They are thinking and will get back to me on the article idea for the frosh supplement | | Gauntlet replied back : They are thinking and will get back to me on the article idea for the frosh supplement |
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| | *From this gel, it looks like trial 1; colonies 1 and 2 might have worked as well as possibly trial one, colonies 3 and 4. The gel looks a little bit slanted however, as the ladder does not perfectly line up on both sides. Regardless, the first two colonies at least look like they may have worked. I will prepare overnight cultures of these colonies as well as colonies 3 and 4. Tomorrow I will isolate plasmids and start with a NottI digest, run this on a gel with my two size controls (J13002-LuxOD47E and BBK LuxOD47E). If this gel looks good, we will send the appropriate colonies for sequencing. I also did restreaks of these colonies. | | *From this gel, it looks like trial 1; colonies 1 and 2 might have worked as well as possibly trial one, colonies 3 and 4. The gel looks a little bit slanted however, as the ladder does not perfectly line up on both sides. Regardless, the first two colonies at least look like they may have worked. I will prepare overnight cultures of these colonies as well as colonies 3 and 4. Tomorrow I will isolate plasmids and start with a NottI digest, run this on a gel with my two size controls (J13002-LuxOD47E and BBK LuxOD47E). If this gel looks good, we will send the appropriate colonies for sequencing. I also did restreaks of these colonies. |
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| - | *Today I also helped Carol do a little but of lab cleanup, filling pipet tip boxes, re-filling tube containers, ethanol bottles, ect. We also went to Invitrogen to get more Polymerase for Carol's stuff. I also helped Carol make restreaks of her glycerol stocks and I saved and edited some pictures for Mandy from the weekend. These pictures will be going up on the Wiki. | + | *Today I also helped Carol do a little but of lab cleanup, filling pipet tip boxes, re-filling tube containers, ethanol bottles, etc. We also went to Invitrogen to get more Polymerase for Carol's stuff. I also helped Carol make restreaks of her glycerol stocks and I saved and edited some pictures for Mandy from the weekend. These pictures will be going up on the Wiki. |
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| - | 9) I had a meeting with Leane Y, she is the communications person at the dept. of biological sciences. She was extrmely happy with our marketing and media progress. I asked her for contact in Alumini and Development offices. Prima will contact the person resonsible for engineering alumini while i will creep on the alumini home page and look for more conatcts. Prima and I will brief the whole team abt the meeting on friday. | + | 9) I had a meeting with Leanne Y, she is the communications person at the dept. of biological sciences. She was extrmely happy with our marketing and media progress. I asked her for contact in Alumini and Development offices. Prima will contact the person resonsible for engineering alumini while i will creep on the alumini home page and look for more conatcts. Prima and I will brief the whole team abt the meeting on friday. |
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| - | 12) I got my US Visa photos ready and payed my deposit for the visa so i am all set (finally) for my appointment on monday july the 20th. | + | 12) I got my US Visa photos ready and payed my deposit for the visa so i am all set (finally) for my appointment on Monday July the 20th. |
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| - | Purpose: To construct the signaling circuit by inserting PQ into B-R-OU-B in AC plasmid, as the pCS26 vector has Kan resistance. This was done by (1) cutting the insert (PQ) with EcoRI and SpeI, and cutting the vector with EcoRI and XbaI, as well as (2) attempting a PQ-B-R-OU-B plasmid switch of colony 12 (see July 14) from AK to AC by cutting the insert and AK plasmid both 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 construct the signaling circuit by inserting PQ into B-R-OU-B in AC plasmid, as the pCS26 vector has Kan resistance. This was done by (1) cutting the insert (PQ) with EcoRI and SpeI, and cutting the vector with EcoRI and XbaI, as well as (2) attempting a PQ-B-R-OU-B plasmid switch of colony 12 (see July 14) from AK to AC by cutting the insert and AK plasmid both with XbaI and PstI. 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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| | * Changing the script for gel electrophoresis to give specific gels back depending on the materials used | | * Changing the script for gel electrophoresis to give specific gels back depending on the materials used |
| | * Adding more conditions to the sequencing machine so that the various products obtained throughout the lab can be analyzed there | | * Adding more conditions to the sequencing machine so that the various products obtained throughout the lab can be analyzed there |
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| | For future needs, I need to take B0034 (RBS) and E0422 (RFP:LVA+Term) out of the 2009 igem distribution plates. B0034 is needed because I need to construct <i>Pqrr4</i>+RBS+GFP:LVA, but the part with GFP:LVA does not contain RBS. E0422 may be needed for characterization. B0034 and E0422 were thus taken out of the plates and transformed into TOP10 cells. They were then grown overnight. Carol promised to take the plates out tomorrow. | | For future needs, I need to take B0034 (RBS) and E0422 (RFP:LVA+Term) out of the 2009 igem distribution plates. B0034 is needed because I need to construct <i>Pqrr4</i>+RBS+GFP:LVA, but the part with GFP:LVA does not contain RBS. E0422 may be needed for characterization. B0034 and E0422 were thus taken out of the plates and transformed into TOP10 cells. They were then grown overnight. Carol promised to take the plates out tomorrow. |
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| | Tomorrow I hope to complete more of DNA extraction (which is taking me longer than it should) . I also have to add variations to it, as right now, it is just the script for a general genomic DNA extraction. These specific variations will be used for the lab missions itself. The general components of lab are just to play around for those who are curious about only particular things, while the specific lab missions have specific goals and completing all 3 will require using each piece of equipment at least once. | | Tomorrow I hope to complete more of DNA extraction (which is taking me longer than it should) . I also have to add variations to it, as right now, it is just the script for a general genomic DNA extraction. These specific variations will be used for the lab missions itself. The general components of lab are just to play around for those who are curious about only particular things, while the specific lab missions have specific goals and completing all 3 will require using each piece of equipment at least once. |
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| | Company 6 : finally got a hold of them, wanted our sponsorship pkg so I sent it to them | | Company 6 : finally got a hold of them, wanted our sponsorship pkg so I sent it to them |
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| - | Company 7 – sent our sponsorship package by mail follow up at the end of july | + | Company 7 – sent our sponsorship package by mail follow up at the end of July |
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| | Company 8 – sent sponsorship package on Monday, called today to follow up – couldn’t get a hold of the CEO so call back tomorrow | | Company 8 – sent sponsorship package on Monday, called today to follow up – couldn’t get a hold of the CEO so call back tomorrow |
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| | Went to visit Leanne Yohemas (Senior Communications Leader – Dept. of Science) – she said she can put up iGEM on the UC twitter and Bio Sci newsletter if we write up an article. She even volunteered to edit and review our articles. We’re thinking of giving her our rough draft in the first week of august. She said she’s willing to be our mentor for promoting iGEM. She also gave us the contact info of the Bio SCI newsletter’s journalist and for the leader/VP of Research in Action (newsletter). Lastly , she gave us the contact info of the person who we should approach for contacts of UC Alumni for funding purposes!! This person is from Development- from the Faculty of Science. I’ll contact her tonight! | | Went to visit Leanne Yohemas (Senior Communications Leader – Dept. of Science) – she said she can put up iGEM on the UC twitter and Bio Sci newsletter if we write up an article. She even volunteered to edit and review our articles. We’re thinking of giving her our rough draft in the first week of august. She said she’s willing to be our mentor for promoting iGEM. She also gave us the contact info of the Bio SCI newsletter’s journalist and for the leader/VP of Research in Action (newsletter). Lastly , she gave us the contact info of the person who we should approach for contacts of UC Alumni for funding purposes!! This person is from Development- from the Faculty of Science. I’ll contact her tonight! |
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| | This didn’t happen the way it was supposed to. I have been building a repository of papers that either focus on quorum sensing (so lots of Bassler and other papers that reference her heavily) or on synthetic biology/biobricking so that I could get a sense of how to structure this. I ended up on a paper by Endy and what looks like 2 of his students that describes their characterisation of the BBa_F2620 part. And once I saw this, that part of modelling took over my life. I sent the paper out to the crew and cc’d the two of you – an explanation of my plan is in my email, but I want to look through it and see what we can easily implement in our work. Because the closer we can mirror this in our circuit characterisation, the better! They seemed to use FACS a lot in their analyses for determining genetic stability. Will we have access to the same technology? Or is there a sneaky way to circumvent it – say, by making a few overnight plates and taking a ratio by sampling the cultures that grow? Anyhow, I shall look at this more tomorrow – I have the paper and the supplement printed, and it would be a good idea to have our experimental procedures ready for this step so that we can hit the ground running when we start. | | This didn’t happen the way it was supposed to. I have been building a repository of papers that either focus on quorum sensing (so lots of Bassler and other papers that reference her heavily) or on synthetic biology/biobricking so that I could get a sense of how to structure this. I ended up on a paper by Endy and what looks like 2 of his students that describes their characterisation of the BBa_F2620 part. And once I saw this, that part of modelling took over my life. I sent the paper out to the crew and cc’d the two of you – an explanation of my plan is in my email, but I want to look through it and see what we can easily implement in our work. Because the closer we can mirror this in our circuit characterisation, the better! They seemed to use FACS a lot in their analyses for determining genetic stability. Will we have access to the same technology? Or is there a sneaky way to circumvent it – say, by making a few overnight plates and taking a ratio by sampling the cultures that grow? Anyhow, I shall look at this more tomorrow – I have the paper and the supplement printed, and it would be a good idea to have our experimental procedures ready for this step so that we can hit the ground running when we start. |
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| | This wasn’t a huge deal – we just had Chinee update the reaction rule chart that we made on Friday, so that the reaction constants that we’re looking up are laid out in front of us on one convenient table. It’s so easy to lose sight of why you’re reading a paper, so this will be useful in helping us keep our goals in mind and in working efficiently. | | This wasn’t a huge deal – we just had Chinee update the reaction rule chart that we made on Friday, so that the reaction constants that we’re looking up are laid out in front of us on one convenient table. It’s so easy to lose sight of why you’re reading a paper, so this will be useful in helping us keep our goals in mind and in working efficiently. |
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| - | I didn’t get around to this and I didn’t want to just start messing around with it without you guys there. If we do end up doing data-based modelling, we will need to have a sense of our delay (time between input and output measurement corresponding to that input). We want this delay to be as low as possible, especially if we’re worrying about protein degradation and time-dependent data (such as response time) – so it would be very prudent to plan our experiments around making that happen. Of course, if the plate takes a long time to read but the reactions are very slow, it won’t be an issue – you just don’t want to lose too much useful information while waiting for the plate measurements to occur.<html> | + | I didn’t get around to this and I didn’t want to just start messing around with it without you guys there. If we do end up doing data-based modelling, we will need to have a sense of our delay (time between input and output measurement corresponding to that input). We want this delay to be as low as possible, especially if we’re worrying about protein degradation and time-dependent data (such as response time) – so it would be very prudent to plan our experiments around making that happen. Of course, if the plate takes a long time to read but the reactions are very slow, it won’t be an issue – you just don’t want to lose too much useful information while waiting for the plate measurements to occur. |
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CAROL
Colony Search
- Final transformation did not result in any colonies. The team has decided to use GFP as a reporter circuit instead since this transformation is not cooperating with us.
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CHINMOYEE
Research
Looked at Papers
Potential :
Found a paper that did deterministic modeling . The paper looks at the autophosphorylation of LuxN ... using as a model for something similar like (LuxPQ) ??? Solution is very complex . If this model will be used---- need to find a simpler way to solve using matlab
More deterministic stuff : Quantitative analysis of protein- protein interactions
Sensitivity amplification in Phosphorylation-Dephosphorylation cycle
Dead End :
Design and sigmalling mechanism of light regulated histidine Kinases
Biological pathway kinetic rate constants are scale invariants
Gauntlet replied back : They are thinking and will get back to me on the article idea for the frosh supplement
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EMILY
Colony PCR to verify Possible Circuit Completion
- Today was an exciting day as my circuit may be finished! I started with a Colony PCR this morning of the J13002-LuxOD47E-B0015 colonies that I transformed yesterday and grew overnight. I ran this Colony PCR on a gel this afternoon. See picture below.
Lanes 1-4 are J13002-LuxOD47E-B0015 trial 1 (treating B0015 as the insert),
Lanes 5-8 are J13002-LuxOD47E-B0015 trial 2 (treating J13002-LuxOD47E as the insert),
Lane 9 is a positive size control J13002-LuxOD47E,
Lane 10 is another positive size control with just BBK LuxOD47E and
Lane 11 is a negative control- ddH2O
- From this gel, it looks like trial 1; colonies 1 and 2 might have worked as well as possibly trial one, colonies 3 and 4. The gel looks a little bit slanted however, as the ladder does not perfectly line up on both sides. Regardless, the first two colonies at least look like they may have worked. I will prepare overnight cultures of these colonies as well as colonies 3 and 4. Tomorrow I will isolate plasmids and start with a NottI digest, run this on a gel with my two size controls (J13002-LuxOD47E and BBK LuxOD47E). If this gel looks good, we will send the appropriate colonies for sequencing. I also did restreaks of these colonies.
- Today I also helped Carol do a little but of lab cleanup, filling pipet tip boxes, re-filling tube containers, ethanol bottles, etc. We also went to Invitrogen to get more Polymerase for Carol's stuff. I also helped Carol make restreaks of her glycerol stocks and I saved and edited some pictures for Mandy from the weekend. These pictures will be going up on the Wiki.
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FAHD
Marketing for July 15th 2009
Today, I concentrated my energies at marketing our iGEM project. Here is what I did today:
1)Called VWR and left him a voicemail. I will get back to him tomorrow
2) I called Charles Rivers Labs, left him a voicemail, i will get back to him tomorrow
3) I called Procter & Gambel Pharma. and inqiured abt our sponsorship package that we had mailed him in June. His assitant told me that he will get back to me next week
4) I called Boheringer Ingleheim Canada and talked abt the Sponsorship package that i had mailed her in June. I left her a voicemail.
5) iGEM was on NUTV. Go to NUTV, Click on shows at the top, Click on Full Frontal at the left top option and watch episode 18. On the bottom of the player it says 'play in DPI quality' so click that option and iGEM is on the second segment of the show (they cut of the Ethics part!!!!!)
6) I emailed a thank you letter to Julie Phillips(News Reporter) and Justin(The programme director) and asked them for a DVD copy and an extended version of the coverage.
7) I e-mailed Rejection Thank you letters to (Conoco Phillips) and (Astellas Pharma)
8) I e-mailed proposals to Alberta Pacific Forest Industries Inc.), (Albian Sands Energy Inc.) and Bietz Resources Ltd.)
9) I had a meeting with Leanne Y, she is the communications person at the dept. of biological sciences. She was extrmely happy with our marketing and media progress. I asked her for contact in Alumini and Development offices. Prima will contact the person resonsible for engineering alumini while i will creep on the alumini home page and look for more conatcts. Prima and I will brief the whole team abt the meeting on friday.
10) I found a couple of group scholarships at the NSERC website whose deadline is Sept 1 and Sept 15. All we have to do is to get someone or ourslves nominate the 2009 iGEM Calgary team for promoting youngsters in the field of science and engineering. The website link is: http://www.nserc-crsng.gc.ca/Prizes-Prix/SciencePromotion-PromotionScience/nomination-nomination_eng.asp. This link will lead you to the sept 1 scholarship while i am still looking for the Sept 15 scholarship.
11) I creeped on AIF's list of partners for contacts which Jen hill told me that she will give them to me.
12) I got my US Visa photos ready and payed my deposit for the visa so i am all set (finally) for my appointment on Monday July the 20th.
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JEREMY
Construction of PQ-B-R-OU-B into AC plasmid
Purpose: To construct the signaling circuit by inserting PQ into B-R-OU-B in AC plasmid, as the pCS26 vector has Kan resistance. This was done by (1) cutting the insert (PQ) with EcoRI and SpeI, and cutting the vector with EcoRI and XbaI, as well as (2) attempting a PQ-B-R-OU-B plasmid switch of colony 12 (see July 14) from AK to AC by cutting the insert and AK plasmid both with XbaI and PstI. 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
Insert Intuitiveness
The main goal I made for myself today was to make the activities I am scripting much more intuitive (especially for first-time second-lifers) then they already are since I believe they seem simple to me only because I was the one who made them. To do this, I have started to create instructions for restriction digest that are not contained within a notecard, which some may be inclined to pass over. So now when the instruction label is clicked (just below the writing – Restriction Digest Start) it will now explain what each section of the activity is for and what you can do (It also lights up each section being talked about to draw the eye), but I would like to improve upon this by adding more detail. They will also get a notecard that explains the restriction digest, ligation and phosphatise treatment aspects of the entire molecular cloning activity.
I also completed the script for a general activity for bacterial transformation so now it will quiz you each time you put a certain object into the tube’s inventory and the questions will relate to the item you added. It then keeps track of the number of questions you answer correctly and if you answered all of the questions right, your competent cells were successful in up taking the DNA. I believe I will make this activity a gradient for now so as long as eight or more questions were answered correctly, the competent cells will survive on their respective plates, just at different levels of growth. I am unsure if I should keep track of each avatars score, though I believe it is possible since I have seen this used other places for different reasons, but it would prevent someone taking credit for someone else’s work. Questions include: What temperature do competent cells need to be stored at?, What does it mean for cells to be competent? etc.
Now I am back to looking at the activities that I began two months ago and am now updating them to suit the lab activities. I have started and will continue tomorrow:
- Cleaning up scripts – getting rid of unnecessary code, making machines work for specific activities
- Renaming items so everything can function together
- Creating instructions for activities such as restriction digest
- Changing the script for gel electrophoresis to give specific gels back depending on the materials used
- Adding more conditions to the sequencing machine so that the various products obtained throughout the lab can be analyzed there
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KEVIN
2. cPCR of Pqrr4+I13500
Since yesterday's cPCR only yielded one possible colony that may contain the circuit of interest, today I performed cPCR on more colonies to see if I can get more to ensure that what I have is what I want. A total of 11 colonies were picked and cPCR was done on then using BBK CP F/R primers. The following is a picture of the gel:
The colonies 2, 3, 7, 9, 10, and 11 seems to have worked. Contamination in negative control, and nothing in positive control. Perhaps the tubes were switched around.
Overnight cultures of yesterday's Colony3, today's colony 2, 3, 7, 9, 10, and 11 were grown for tomorrow's restriction digest.
2. Transformation of B0034 and E0422
For future needs, I need to take B0034 (RBS) and E0422 (RFP:LVA+Term) out of the 2009 igem distribution plates. B0034 is needed because I need to construct Pqrr4+RBS+GFP:LVA, but the part with GFP:LVA does not contain RBS. E0422 may be needed for characterization. B0034 and E0422 were thus taken out of the plates and transformed into TOP10 cells. They were then grown overnight. Carol promised to take the plates out tomorrow.
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MANDY
DNA Extraction in Second Life
Today, I continued on with scripting DNA extraction (and still am not done :( ). I kept the centrifugation at each step, so I had to script the centrifuge to work with different ‘solutions’ and make the test tubes and other components move to and from the centrifuge smoothly. I wasn’t able to script the centrifuge door to close when it was being used, so it has to be manually done by the user. I rewrote the addition of buffer part so that centrifugation is required after each step. I had to rewrite most of the scripting I did yesterday due to the centrifuge part, but right now it all works up to the steps that require addition of buffer, which I still have to finish.
The Linden in charge of education hasn’t replied to our message yet, so if he doesn’t by the end of the week, Stefan and I will visit him during his office hours on Tuesday.
Tomorrow I hope to complete more of DNA extraction (which is taking me longer than it should) . I also have to add variations to it, as right now, it is just the script for a general genomic DNA extraction. These specific variations will be used for the lab missions itself. The general components of lab are just to play around for those who are curious about only particular things, while the specific lab missions have specific goals and completing all 3 will require using each piece of equipment at least once.
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PATRICK
Names and Commas
Retrospective Notebook: This entry was not written on this day, but derived later from working notes I made that day.
Began working on putting the important DNA part information into the object's names. Briefly toyed with the idea of making all complexes (be they protein, DNA, small cofactors, or any combination of the above) user definable, before determining that this would be extremely difficult. Began thinking about how the Biobricker and its construction feature would work exactly, what types of information would each would need from the other.
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PRIMA
Marketing, media and advertisements
For marketing:
I called these companies again today:
Company 1 – filled out online application for sponsorship so follow up phone call July 20th
BioAlberta –I called to follow up but he wasn’t there so I left a msg and sent him an email.
Company 3 – they received our package and looking through , follow up July 24
Company 4&5 : follow up july 24 – received our package
Company 6 : finally got a hold of them, wanted our sponsorship pkg so I sent it to them
Company 7 – sent our sponsorship package by mail follow up at the end of July
Company 8 – sent sponsorship package on Monday, called today to follow up – couldn’t get a hold of the CEO so call back tomorrow
Company 9, 10, 11 : received sponsorship package so follow up July 20
Found 10 more biotech companies online who we could contact. So I assigned and distributed the new list of companies to the rest of the marketing team! :D
I wrote up 3 short articles for the July newsletter (Lethbridge workshop, 3rd bake sale, Sponsor of the month).
Went to visit Leanne Yohemas (Senior Communications Leader – Dept. of Science) – she said she can put up iGEM on the UC twitter and Bio Sci newsletter if we write up an article. She even volunteered to edit and review our articles. We’re thinking of giving her our rough draft in the first week of august. She said she’s willing to be our mentor for promoting iGEM. She also gave us the contact info of the Bio SCI newsletter’s journalist and for the leader/VP of Research in Action (newsletter). Lastly , she gave us the contact info of the person who we should approach for contacts of UC Alumni for funding purposes!! This person is from Development- from the Faculty of Science. I’ll contact her tonight!
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STEFAN
Variety of things to do
so what am I doing today?
- Fixing the Synthetic Kingdom video. Technical problems plagued my blog post and thus, took forever to do. Apparently, it also has no sound so I have to find a way to fix that tonight. I think the problem is on youtube's end of things.
- Acting as an EXTERIOR decorator for the Synthetic Kingdom. The purpose of this is to make look nicer and not retina-burning ugly. People would feel better exploring it and it would provide more incentive to stay there and see all it has to offer.
- Building a new Endoplasmic Reticulum for my cell. I spied on iGEM Washington's Second Life stuff and found a better way to build it (they stole it from somewhere! tsk tsk).
- Trying to read and take notes on another ethics paper because the meeting will be coming up soon. Also, I wanted to look into the Ethics Approval to see if we can interview people without breaking the law.
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VICKI
Technical report writing
This didn’t happen the way it was supposed to. I have been building a repository of papers that either focus on quorum sensing (so lots of Bassler and other papers that reference her heavily) or on synthetic biology/biobricking so that I could get a sense of how to structure this. I ended up on a paper by Endy and what looks like 2 of his students that describes their characterisation of the BBa_F2620 part. And once I saw this, that part of modelling took over my life. I sent the paper out to the crew and cc’d the two of you – an explanation of my plan is in my email, but I want to look through it and see what we can easily implement in our work. Because the closer we can mirror this in our circuit characterisation, the better! They seemed to use FACS a lot in their analyses for determining genetic stability. Will we have access to the same technology? Or is there a sneaky way to circumvent it – say, by making a few overnight plates and taking a ratio by sampling the cultures that grow? Anyhow, I shall look at this more tomorrow – I have the paper and the supplement printed, and it would be a good idea to have our experimental procedures ready for this step so that we can hit the ground running when we start.
Modelling effort coordination
This wasn’t a huge deal – we just had Chinee update the reaction rule chart that we made on Friday, so that the reaction constants that we’re looking up are laid out in front of us on one convenient table. It’s so easy to lose sight of why you’re reading a paper, so this will be useful in helping us keep our goals in mind and in working efficiently.
Plate reader
I didn’t get around to this and I didn’t want to just start messing around with it without you guys there. If we do end up doing data-based modelling, we will need to have a sense of our delay (time between input and output measurement corresponding to that input). We want this delay to be as low as possible, especially if we’re worrying about protein degradation and time-dependent data (such as response time) – so it would be very prudent to plan our experiments around making that happen. Of course, if the plate takes a long time to read but the reactions are very slow, it won’t be an issue – you just don’t want to lose too much useful information while waiting for the plate measurements to occur.
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