PROJECT

RESULTS

Day 25 - Monday, July 18th 2011

The ligated transformations that did not grow on Friday appeared to grow in the incubator over the weeekend. There were both red and white colonies on each plate.
We ran digestions of our RBS ligation minipreps out on a gel. We wanted to make sure the ligated part was responsible for the transformation and not some uncut backbone. Both gels showed almost all the colonies we overnighted had bands at about 700 base pairs, a good sign that our ligated part was in the backbone.
Cut our genomic LasR with all of the biobrick restriction enzymes to confirm our restriction site analysis. The results generally matched what we expected.
Miniprepped our new RBS part and another round of ligated transformants.
Started assembling promoters onto the previously ligated RBS -> Reporter and RBS -> Receptor ligations. We are doing the 3A assembly this afternoon and will attempt our 2A method tomorrow.
Sent samples of many of our parts and ligations in for sequencing.

Day 26 - Tuesday, July 19h 2011

Began transforming the parts we ligated yesterday with the 3A assembly. We also transformed a positive control using only 0.5ng of DNA so we get fewer colonies and can make a more accurate measurement of our competency.
Miniprepped the overnights of the new RBS and the old RBS-part ligations that we don’t already have glycerol stocks of.
Began a test run of PCRing our miniprepped parts with the biobrick sequencing primers. This would give us a quick and easy way to get tons of DNA.
Started a 2A assembly of RBS -> Repoters and RBS -> Receptors
Started a 2A assembly of Promoters -> [RBS -> Receptors and Repoters]
Researched natural autoinducer concentrations in Pseudomonas so we would know how much autoinducer to order.

Day 27 - Wednesday, July 20th 2011

The transformations did not grow in time to overnight last night, so we will overnight them later today. All of the plates now have colonies.
We miniprepped the overnights of the last remaining part (RBS -> RhlR) that we don’t have a glycerol stock of.
We transformed our 2A ligations from yesterday
We attempted another round of PCR, as our gel yesterday had no bands. We are going to try using more DNA this time.

Day 28 - Thursday, July 21st 2011

Miniprepped all of our 2A assembly ligations. Since many plates had both red and white colonies, we made overnights of both. We suspect the red colonies are the backbone, but want to confirm this by running a gel.
Started a 3A ligation with the new RBS.
Mapped out our current progress in the project and a plan for next week.
Ligated several promoters with RBS34 and parts
PCRed our miniprepped parts so we have more DNA in storage.

Day 29 - Friday, July 22nd 2011

Miniprepped the latest overnights of our 2A ligations.
Digested all of our 3 part ligations with X and S so that we can run them on a gel next week to see exactly what is in all of our colonies.
Got our sequencing data for the first two registry parts, still need to analyze.
Transformed a round of RBS34 ligations, overnights will be started on Sunday so we can miniprep Monday.