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| - | cPCR of <i>Pqrr4</i>+I13500 | + | 2. cPCR of <i>Pqrr4</i>+I13500 |
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| | 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. | | 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. | | Overnight cultures of yesterday's Colony3, today's colony 2, 3, 7, 9, 10, and 11 were grown for tomorrow's restriction digest. |
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| | + | 2. Transformation of B0034 and E0422 |
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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. |
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CAROL
Descriptive Title of What You’re Doing
WIKI CODING HERE
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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.
- Analysis:
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, 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.
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FAHD
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IMAN
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JAMIE
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JEREMY
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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
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PATRICK
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PRIMA
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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
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