Team:UIUC-Illinois/Notebook/MeetingNotes

From 2012.igem.org

(Difference between revisions)
Line 2,138: Line 2,138:
<div id="meet30" style="display:none">
<div id="meet30" style="display:none">
<textarea readonly rows="30" wrap="soft">
<textarea readonly rows="30" wrap="soft">
-
 
+
Monday, 7/9/12
 +
- Adi - meeting with Joe this week to look over entrepreneurship executive summary, are being sponsored by SARSTEDT (box of supplies came today), emailed thank yous to sarstedt and gold biotech. Should contact Octochem about further sponsorship.
 +
- Divya, Washington characterization - did the M9 media for alkane production, for publicity is writing an email to IMSA, Asha took pics for blog, Divya will email Flickr password out. Will talk to Isiah about tshirts. After actually obtaining alkanes, will write up the characterization with isiah. Followed a bunch of people on twitter (37), and currently have 20 followers. Also need to make a new trifold for Quad day and E-night. Add any other publicity ideas to the google doc.
 +
- Cara, PHAT - have designed sequencing primers but not ordered them yet (wants to try using VR and VF), might need to design another set of sequencing primers to account for 100 bp mess up at the start of every sequence. UPenn is going to ship genetic info on rotobacter at sometime next week because the rotobacter from Dr. Metcalf isn’t working. Is talking to someone in MMG about high performance thin plate chromatography. Have glycerol stocks of BM3s in expression vector. Get things characterized before biobricking (plus, biobricking primers aren’t in right now). Would like to buy malonyl-CoA for $80, but haven’t talked to Courtney about it yet.
 +
- Bob, webmaster - redoing PCR for 4CL:STS and will run gel later, wiki will be changed so that the lab notebook is separated among the PUF and PHAT projects. Have bullet points and pictures, doesn’t need to be very intense. Keep it simple and clear. Each day has a data section with numbers or pictures divided into the PUF and PHAT projects. Can also have tabs and drop down menus. Also uploaded most recent minutes. Will also add Sarstedt to the sponsors page.
 +
- Anthony, PUF - did all steps from PCR to transformation of WT PUF. The transformation didnt work so have started redoing it.
 +
- Asha, PUF - working on re-doing the eYFP in protet constructs (both the control binding site and the PUF binding site.) Did PCR today, and needs to run the gel later. Then will religate, transform, grow, do colony PCR, send to sequencing and begin characterization. Eventually, will clone in the WT PUF in pBAD to the cells. Also making plate (10 LB, 25 Amp, 25 Amp and Cm) and making more pBAD. Will contact local science camps about presentations/assisting in lab for outreach.
 +
- Angela - after doing digestion, you don’t always have to do gel purification, can just do PCR cleanup. Because alot of these runs are low copy number, so gel purification yields low numbers. Did tons of PCR for PUF and most worked. No pBAD worked in the digestion, so got more pBAD from Asha and will hopefully have finished digestion/ligation/transformation by tomorrow. Will then send in for sequencing.
 +
- Uros - trying to get the pETDUET to work with split GFP, but hasn’t really worked. the pETDUET plasmid is low copy number and so we have low yield and have a really faint band with split GFP. If we sent the split GFP tethered to PUF to sequencing on Friday, we would get it at the end of July. It’s expensive and takes a while but we know that we would have it. If we did it by ourselves it would be difficult, and take an undetermined amount of time.
</textarea>
</textarea>
</div>
</div>
 +
<div id="meet31" style="display:none">
<div id="meet31" style="display:none">
<textarea readonly rows="30" wrap="soft">
<textarea readonly rows="30" wrap="soft">
-
 
+
Tuesday, 7/10/12
 +
- Divya, washington characterization - still waiting for the stuff to grow, will email Isiah about further updates, need to talk to Jin about GCMS on Thursday. helped Asha plate some stuff, will make plates tomorrow.
 +
- Anthony - looked a PCR gel. only 1 of 3 worked, but it was a lot of yield, so it will last a couple of rounds of digestion/ligation. So the digest/ligate will occur today and be finished tomorrow.
 +
- Asha -YFP PCR worked. looked at other YFP colonies under UV light and none of them glowed =( so it’s a good thing we’re re-ligating! ruined 70 plates... sorry! =(
 +
- Angela - did ligation/transformation and will see if it worked later tonight. designing primers for primer extension (backup plan). Being trained in Rao’s lab to use sewing and use the florescent meter to measure the mcherry and venus for the reporter genes. Asking to be trained for mathematical modeling for the output of our project.
 +
- Uros, scaffold and tethering - working to tether PUF and GFP. Read the UNC paper (draft) on how they tethered, but will need more info because there is no supplemental info or data. Also need to send some emails out and continue reading up on it. Wants to talk to Todd about tethering and the low copy plasmid. Ligations didnt work and transformations didnt work =(
</textarea>
</textarea>
</div>
</div>
 +
<div id="meet32" style="display:none">
<div id="meet32" style="display:none">
<textarea readonly rows="30" wrap="soft">
<textarea readonly rows="30" wrap="soft">
-
 
+
Wednesday, 7/11/12
 +
- Asha, PUF - yesterday digested eYFP and protet and got bands of the correct length. Ran ligation for 7 hours using a 10:1 ratio of insert:vector. Transformed today, Bob and Cara plated for me. Will look at plates tomorrow. If colonies, will look at them under the UV light to see if they glow!
 +
- Divya, made CM, Amp, and Amp+CM plates for Asha, talked to Jin about GCSM: work with Quan Lee (?), Isiah will pick up alkane production tonight, will resuspend tomorrow in EtOAc to extract
 +
-Bob, put Starssted on the sponsor’s section, and worked
 +
-Angela:
 +
Constructed primers using primer extension method for the entire PUF project constructs. Primers for cloning of YFP to ProTet and wild type PUF to pBAD. To be more precise the method is termed CPCA used for cloning. The benefits are that it doesn’t need any ligation or gel purification. Once the primers arrive, probably Friday, we can try the PCR and check whether the insert is present or not by check primers. Check primers were ordered today as well. Wild type PUF need another set of sequencing primers. Orders have actually not been placed yet, it depends on Courtney. CPAC method doesn’t leave any scars in the cloning and no gel purification is required, only PCR and gel running is required. Perhaps the method will be taught sometime later in the week to the whole group. One modification is that instead of using E0030, Venus will be used. PSB1C3 will be arriving on Friday. Transformation after ligation was done today, colonies will be seen tomorrow. Starting another set of regular cloning in case CPAC doesn’t work, from digestion/ligation/transformation.
 +
-Adi:
 +
Alex will be arriving this weekend. The entrepreneurship wiki should be done in approximately 2 weeks. Adi will e-mail Mark from Octochem about a chemical needed for the PHAT project.
 +
-Anthony:
 +
Digested, extracted, and ligated PCR of pBAD and wild type PUF PCR. Transformed today and will plate, hopefully there will be colonies tomorrow.
 +
-Uros:
 +
Continued to investigate how to tether PUF to a correct split-GFP sequence. Asked Angela for further information on the UNC paper in order to receive more information where on PUF was the tether placed. Proceeded to e-mail the graduate student who helped collaborate and give advice for the RNA Scaffold synthesis about the split-GFP sequence used in the paper (Pam Silver paper). E-mailed Dr. Bourde, the principal investigator of the lab which was referenced in the Pam Silver paper, concerning split-GFP work for either the sequence or possible part shipment. E-mailed Todd in order to get more information or suggestions on where to head with the tethering project. Also, Todd might offer insight whether it is still feasible to produce the construct ourselves before August and so we could save money before synthesis. Been trying to find a trustworthy source for the split-GFP sequence and have found two sources. One of them being from a past iGEM team and the other from a research article. For Friday an update will be presented to the advisors along with a list of options which can be pursued, one of which will be decided upon collectively. An e-mail was forwarded from Courtney, sent by IDT, that the RNA scaffold gene is still in production and everything is being done so it could be properly made. Any other product ordered along with the scaffold should be asked to be sent immediately due to the unpredictable completion time of it.
 +
-Cara/Bob:
 +
Cara and Bob made smaller aliquots of freshly made LB media and put them into the 50mL conical tubes. The contents of the conical tubes should be transferred into small glass containers and the tubes autoclaved/reused. Gloves arrived today, needed materials need to be updated and immediately communicated to Courtney. Rafferdy got back to Cara and has TLC plates for her to use. TLC standards worked yesterday, solvent used was 85:15:3 chloroform, methanol, acetic acid. Reverse phase was tried as well. 10mL of solvent was used, 250mL of solvent was made which should last a while. All reagents were received from MMG. Gels were ran to check whether PCR worked last night, PCR might have worked. The gels need to be extracted and sequenced in order to check whether it worked. IDT will be e-mailed/called to separate the orders of scaffold and primers due to delay of the scaffold production. 5mg of piceattanol and 100mg of resveratrol are in the inventory. All piceattanol contents were resuspended in ethanol. Resveratrol will be diluted as well in some solvent. Starting fermentation is the next step. Bob plated some things for Asha after transformation. Sponsors were also added to the Wiki and the lab notebook section was updated. Meeting with Melissa will be held by the end of the week. WashU credited UIUC coding done by Bob for the Wiki!!
</textarea>
</textarea>
</div>
</div>
 +
<div id="meet33" style="display:none">
<div id="meet33" style="display:none">
<textarea readonly rows="30" wrap="soft">
<textarea readonly rows="30" wrap="soft">
-
 
+
Thursday, 7/12/12
 +
- Divya - Isiah got the cultures out. Talked to Jin and it turns out that we actually need to wait until the alkanes arrive to use them as a standard for GCMS. Talked to the GCMS lady about conditions. Divya will look up the info and email it to the lady to create the standard curve. Either tomorrow or monday results will occur. On next Monday, will begin helping with cloning.
 +
- Asha - transformation of YFP ligation didn’t work, so replated. Also another ligation with only a 5:1 insert:vector ratio. Also went to the art department to ask about filming equipment for human practices. The checkout counter wasn’t open, but got an email to contact.
 +
- Anthony - failed transformation from yesterday so replated. Also transformed again with a different ligation. Among 4 plates hopefully there will be results! =)
 +
- Bob - ran a gel of Cara’s PHAT optimized PCR. Started a digestion of it. Working on wiki, more specifically putting all of the lab notes from the google doc on the wiki in nice little tables. Will also contact Melissa.
 +
- Cara - did gel purification of the PHAT optimized PCR. The band is not the right length, but the PCR has consistently yielded this band. Therefore purified it and wants to find what it is. The suspicion is that the primers and PCR are working and the part is messed up. Digesting the vector or sequencing the part are good things to do at this point. Will type up a protocol from Brad and send it to everyone about growing things in a controlled environment for PHAT.
 +
- Uros - talked to Todd in detail about the tethered PUF to CFP construct ourselves. He thinks that we can do it ourselves and complete it before August. The plan involves making alot of primers (angela will help). In parallel will construct a plasmid with the CFP parts so that we can have a control to see how the CFP works. have to check if CFP is in the kit and if we can just PCR it. Still the debate: do we synthesize or do it ourselves? it’s the problem of time and money. will sit down and create a very detailed plan with Todd tomorrow.
 +
- Adi - been working on the wiki, Alex will come down on Saturday and everyone should come to lunch to meet him and discuss
 +
- Angela - everyone needs to create a powerpoint of work and data to present at the advisor’s meeting tomorrow. Having problems with comp cells because transformations aren’t working. So is going to take some iGEM comp cells and try the same ligation transformations. It that doesn’t work will then transform with boughten comp cells. Prepared more ligations so will continuously ligate and transform until results. Finished the primers for primer extension and checked with Kori about making a standard protocol. Next week the primers will arrive and will start cloning the modular PUF reporters via primer extension.
</textarea>
</textarea>
</div>
</div>
 +
<div id="meet34" style="display:none">
<div id="meet34" style="display:none">
<textarea readonly rows="30" wrap="soft">
<textarea readonly rows="30" wrap="soft">

Revision as of 19:12, 2 October 2012

Header

Protocols

Meeting Notes

Pre-April

  • 2/17/12
  • 2/19/12
  • 2/26/12
  • 3/4/12
  • 3/6/12
  • 3/7/12
  • 3/11/12
  • 3/25/12
  • 3/27/12
  • April

  • 4/1/12
  • 4/3/12
  • 4/4/12
  • 4/8/12
  • 4/14/12
  • 4/15/12
  • 4/17/12
  • 4/22/12
  • 4/24/12
  • May-July

  • 5/1/12
  • 5/25/12
  • 5/29/12
  • 6/1/12
  • 6/11/12
  • 6/12/12
  • 6/18/12
  • 6/19/12
  • 6/20/12
  • 6/21/12
  • July-August

  • 7/5/12
  • 7/6/12
  • 7/10/12
  • 7/11/12
  • 7/12/12
  • 7/16/12
  • 7/17/12
  • 7/19/12
  • 7/20/12
  • 7/25/12
  • 7/30/12
  • August-Jamboree

  • 8/6/12
  • 8/10/12
  • 8/24/12
  • 8/25/12
  • 8/31/12
  • 9/15/12
  • Select a date to read respective meeting notes.


    Retrieved from "http://2012.igem.org/Team:UIUC-Illinois/Notebook/MeetingNotes"