Team:Northwestern/Notebook

Notebook

Weekly Accomplishments

Brainstorming April-June 2010

Week1 6/13/10-6/19/10

Week2 6/20/10-6/26/10

Week3 6/27/10-7/3/10

Week4 7/4/10-7/10/10

Week5 7/11/10-7/17/10

Week6 7/18/10-7/24/10

Week7 7/25/10-7/31/10

Week8 8/1/10-8/7/10

Week9 8/8/10-8/14/10

Week10 8/15/10-8/21/10

Week11 8/22/10-8/28/10

Week12 8/29/10-9/4/10

Week13 9/5/10-9/11/10

Week14 9/12/10-9/18/10

Week15 9/19/10-9/25/10

Week16 9/26/10-10/2/10

Week17 10/3/10-10/9/10

Week18 10/10/10-10/16/10

Week19 10/17/10-10/23/10

Week20 10/24/10-10/30/10

Daily Accomplishments

10/3/10

Timi miniprepped the CP-LacPI-GFP cultures and digested them with E/P. She then poured a big and a small gel.

Kevin came in and ran the digested CP-LacPI-GFP, digested CP-LacPI with digested Cp and digested LacPI parts.

10/4/10

Timi and Kevin decided to take a day off :)

10/5/10

Timi and Kevin came in to discuss why their gels were running strangely. We believe there is something wrong with our buffer, so we are swapping everything out with a fresh batch of Prof. Mordacq's 0.5x TBE buffer. Timi poured a large gel that we will be using tomorrow.

10/6/10

10/7/10

Timi- took part 2-17F out of the DNA kit. Transformed into T10 cells to grow overnight.

10/8/10

Kevin- ran gel of the CP-LacPI parts. Gel results are still inconclusive. Also put in an overnight culture of the 2-17F part into the incubator for Timi.

10/9/10

Timi- Miniprepped the 2-17F part. Digested the DNA with E/P and ligated the 2-17F GFP part into Chlor and Tet backbone.

10/10/10

Timi- Transformed and plated part 2-17F onto Chlor and Tet plates. Will be ready for miniprepping tomorrow.

10/11/10

10/12/10

Matt - digested LacP-RBS with SpeI in preparation for standard assembly with ES digested CHS3

10/13/10

10/14/10

Kevin transformed T10 cells with CP-LacPI parts that we did not have much digest of left. He plated them onto TET plates.

10/15/10

Kevin took out the CP-LacPI plates and put them into the 4 degree for Timi. Timi inoculated overnight cultures at 7pm.

10/16/10

Team meeting at 12pm to work on our wiki. We made the discouraging discovery that our CHS3 DNA that Ben had worked super hard for actually contained XbaI restriction sites. This complicates our assembly and might jeopardize the success of our project. We will talk to the advisors about this to see if things can still work out.

Timi and Kevin miniprepped the CP-LacPI inoculations, digested them with E&P, and ligated them to GFP.

10/17/10

Timi phosphotase-treated the newly digested chlor backbone and ligated a second set of CP-LacPI. Kevin transformed and plated this new set of phosophotase treated Cp-LacPI.

Kevin inoculated the first set of non-phosphotase treated CP-LacPI inoculations.

10/18/10

Matt miniprepped 31GFP, 21GFP (old), and 32GFP (old). He digested the miniprepped DNA with E/P for Kevin to run on a gel. He also reinoculated all 9 CP-LacPI combinations.

Matt inoculated the phosophotase-treated ligations at 9pm.

10/19/10

Plan: Timi miniprepped the phosphotase-treated CP-LacPI and ran them on a gel. But Mordacq's class was using the thermocycler so Timi just sat and read.

Plate reader fun with Kevin @ 3pm.

Sean - ran CHS3(pMAL+SDM) digest, pMAL digest, and ligation on a gel -> CHS3 exhibited incorrect bands (there are 2, when there should be 1) ligation showed nothing -> plan to run CHS3 undigested and rerun the above 3.

10/20/10

Timi and Kevin ligated the old 212, 312, and 322 parts into a phosophotase-treated Chlor backbone for submission into the registry. They transformed and plated for inoculation tomorrow night.

They inoculated cultures for the plate reader.

10/21/10

Timi/Kevin: Early morning plate reader protocol entering.

Weekly lab meeting with advisors.

10/22/10

PCR'ed out CHS3 with pMAL ends and Gel extracted

Kevin was in lab until 3am to see if any of the CP-LacPI parts were ligated to GFP. He ran multiple gels and sent the results to Prof. Leonard. Unfortunately, we do not the have the construct ready. Thus, we will likely not be able to characterize a CP-LacPI part by Wednesday. However, we plan on working to characterize one by the Jamboree.

10/23/10

Timi miniprepped the overnights for DNA that will be submitted the registry! We will be sending in parts http://partsregistry.org/Part:BBa_K418003 BBa_K418003, http://partsregistry.org/Part:BBa_K418005 BBa_K418005 and http://partsregistry.org/Part:BBa_K418006 BBa_K418006 on Monday afternoon via Fedex. Yay!!

Timi and Kevin retransformed the confirmed CP-LacPI parts (CP2-LacPI1, CP3-LacPI1 and CP3-LacPI2) from earlier in the summer into ChiA competent cells. The cells were plated onto Chlor plates. We will be continuing our work to hook them up to GFP over the subsequent weeks. Hopefully, we will have all of our work done by the Jamboree! Wish us luck!

10/24/10

Timi took out the CP-LacPI plates into the morning and stored them in the cold room.

Kevin inoculated overnight cultures and put them in to grow.

10/25/10

Kevin miniprepped the CP-LacPI cultures and digested them.

10/26/10

Timi treated Tet backbone with phosphatase for more efficient 3A assemblies.

Timi ligated the CP-LacPI digests with GFP into a Tet backbone. They were transformed into ChiA competent cells and plated onto Tet plates.

10/27/10

Kevin inoculated overnight cultures of the CP-LacPI-GFP colonies. They will be miniprepped/digested tomorrow to be screen on an agarose gel. After we successfully obtain the part, we will characterize the CP-LacPI-GFP constructs that we created to find the weakest and strongest constitutive promoter. We will be using a plate reader to characterize the parts.

WIKI FREEZE at 11:59PM ET. Any further work will be documented elsewhere.