Week of July 10 - July 16
Monday morning, the preservation protocol setup was begun. We prepared medium, plated negative controls, and inoculated wild-type E. coli to grow overnight. Additionally, we digested and purified gBlock 4 for the TodX transporter, performed a PCR on gBlocks 1-3, ran a gel on colony PCR and Gibson and did a PCR check of the Gibson. We also created an LB-agar solution, inoculated LB to prepare for the MIC50 assay, and tested electrocompetent cells’ transformation efficiency. In the dry lab, we had Skype calls with Teams Peshawar and Valencia, contacted Team Toulouse to follow up with them about the preservation protocol, and read over the InterLab Study 2017 documentation to prepare for plate reading.
On Tuesday, our former president Mark dropped in for the day to work on parts of the mathematical modeling. We continued our preparations for the preservation protocol. We found a positive result on our negative control, and so boiled the medium to sterilize it. We also performed PCR for the gBlocks, ran gels on the gBlocks, and excised the correct bands. In the afternoon, we had a Skype call with Team Rose-Hulman and published a note on the Experiment Campaign.
Wednesday, we started the day by finding another positive on our negative control, and performed another sterilization of the medium. We also began using the flex station to create a standard fluorescence curve of NADH, which failed, and set up the benzene growth inhibition assay. Also in the wet lab, we made a solution of Tris-HCl, made a Gibson Master-Mix, performed gibson assembly on the upper and lower operons, digested BnzABCD and TodX, ran gels on the digestions, prepared PCR for gBlocks and miniprepped gBlock 4. We continued working on the mathematical model by incorporating the FBA/FVA connection to glucose uptake and cell growth in tandem with benzene degradation. We also called the United Steelworkers union to speak to them about their opinions and concerns about the project and ordered our universal promoters through IDT.
On Thursday, we continued our mathematical modeling by creating graphs of data obtained by the code and implementing a model of cell growth coupled with benzene degradation for cell groups. In the wet lab, we ran and excised a gel of gBlock3 PCR products, performed a Gibson PCR check for the upper and lower operons, ran a gel of the Gibson PCR, performed a TodX gBlock 4 digest, started a transport assay for growth and induction to collect cells, and continued the benzene growth inhibition assay.
Friday, we obtained a ventilator from Dr. Zhongming Liu’s lab for the lungs being received on Tuesday. We made some other preparations for the lungs and had a meeting with Lafayette’s children’s
science museum, Imagination Station, about hosting an outreach event. We performed PCR and gels on the upper and lower operons, Gibson assembly of the operons, and gBlocks 1 and 2. We also
continued the benzene growth inhibition protocol, continued the transport assay for the cells, and digested gBlock 4. In the dry lab, we finished a new draft for the wiki homepage!