{{:Team:Northwestern/Templates/trial}}

{{:Team:Northwestern/Templates/trial}}

*The transformations from the weekend had some very weird results. They were all red despite our attempts to ligate GFP. We think the issues is with our Kan backbones (uncut backbones may have transformed the bacteria)

*The transformations from the weekend had some very weird results. They were all red despite our attempts to ligate GFP. We think the issues is with our Kan backbones (uncut backbones may have transformed the bacteria)

*We began another batch of competent cells, using a slightly different procedure from a molecular cloning manual.

*We began another batch of competent cells, using a slightly different procedure from a molecular cloning manual.

*Ran the PCR products out on a gel to see if the process was successful

*Ran the PCR products out on a gel to see if the process was successful

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*Our SOB broth was contaminated! The test tube we grew overnight containing just broth was still cloudly in the morning. As a result, our competent cell procedure will have to be restarted and new SOB will have to be made.  

*Our SOB broth was contaminated! The test tube we grew overnight containing just broth was still cloudly in the morning. As a result, our competent cell procedure will have to be restarted and new SOB will have to be made.  

*Because the overnights had an antibiotic, we decided to miniprep anyway despite the contamination.  

*Because the overnights had an antibiotic, we decided to miniprep anyway despite the contamination.  

*We started a digest of our RBS parts and our eventual downstream parts so that we can ligate them together via a modified 1A assembly. The procedure leaves the promoter in with its backbone, making gel extraction possible.  

*We started a digest of our RBS parts and our eventual downstream parts so that we can ligate them together via a modified 1A assembly. The procedure leaves the promoter in with its backbone, making gel extraction possible.  

*We also again attempted to digest and ligate our promoters together with our reporters. This will allow us to test if our promoters are working.

*We also again attempted to digest and ligate our promoters together with our reporters. This will allow us to test if our promoters are working.

*Ran the PCR products from yesterday on a gel. This time, neither gene appeared to be successfully amplified. All we saw were two small, unidentified bands.  

*Ran the PCR products from yesterday on a gel. This time, neither gene appeared to be successfully amplified. All we saw were two small, unidentified bands.  

*Competed cells procedure restarted with fresh, uncontaminated SOB

*Competed cells procedure restarted with fresh, uncontaminated SOB

*We also started a modified 3A Assembly with similar parts. This procedure requires gel extraction, which took longer, so the ligation was left to run overnight.  

*We also started a modified 3A Assembly with similar parts. This procedure requires gel extraction, which took longer, so the ligation was left to run overnight.  

*We ordered the sequencing primers and the primers that will allow us to extract the promoters from the genome.  

*We ordered the sequencing primers and the primers that will allow us to extract the promoters from the genome.  

*Miniprepped 28 overnights of all of our parts and backbones. The DNA concentrations were generally very good, among the best that we’ve had so far.  

*Miniprepped 28 overnights of all of our parts and backbones. The DNA concentrations were generally very good, among the best that we’ve had so far.  

*Many of our ligated transformations grew! There were colonies on almost every plate. This is a good sign, although we will have to sequence and/or run a gel to make sure the cells were transformed with the correct plasmid.  

*Many of our ligated transformations grew! There were colonies on almost every plate. This is a good sign, although we will have to sequence and/or run a gel to make sure the cells were transformed with the correct plasmid.  

*Made new Tet and Amp plates

*Made new Tet and Amp plates

*Decided to start overnights from the successful transformations, but only for the parts with Ribosome Binding Sites. Our advisors definitely think this will improve efficiency.  

*Decided to start overnights from the successful transformations, but only for the parts with Ribosome Binding Sites. Our advisors definitely think this will improve efficiency.  

*Miniprepped the overnights of our ligated RBS parts.  

*Miniprepped the overnights of our ligated RBS parts.  

*Started a PCR reaction with our new primers to hopefully extract Las and Rhl sensitive promoters directly from the pseudomonas genome. We also attempted to extract RhlR again.  

*Started a PCR reaction with our new primers to hopefully extract Las and Rhl sensitive promoters directly from the pseudomonas genome. We also attempted to extract RhlR again.  

*Cut our ligated and miniprepped overnights with different restriction enzymes and ran it on a gel to confirm the presence of two P restriction sites.

*Cut our ligated and miniprepped overnights with different restriction enzymes and ran it on a gel to confirm the presence of two P restriction sites.