Team:Queens Canada/Notebook/Week9


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Notebook - Week 9

Protocol Content

Monday June 25

In the morning, Phillip and Victor did a PCR and gel of fliC1 using a newly grown liquid culture of BL21 and trying different methods to isolate the template DNA better. The gel still turned out blank. What will it take to see that little white band show up ~700 kb on the dark imposing gel landscape? The rest of us did some research on new primers and zinc finger nucleases. In the evening, the members of SynthetiQ convened. We made a timeline, wish list of concepts we would like to illustrate, and crossed our fingers for an artist-in-residence.

Tuesday June 26

In the morning, we examined a potpourri of research topics that included zinc finger nucleases, converting the E. coli fliC sequence to BB-2 standard, analysis of the S. typhimurium fliC sequence, and genomic DNA isolation protocols. In the lab, David and Victor resuspended the fliC second constant domain we got synthesized, performed heat shock transformation, and grew the transformed bacteria on three ampicillin plates. Kevin had a meeting with Dr. Steven Smith and came back with an exciting new prospect: using cohesins and dockerins in our chimeric flagella. These proteins are part of a multi-enzyme complex called the cellulosome that some anaerobic bacteria use to degrade cellulose. This might just be the perfect application for our flagellar scaffold that we have been chasing after these past two months. In the afternoon, we all plunged right into this research topic. Rather than forge ahead with another haphazard colony PCR on a very limited supply of Hot Start polymerase, we have decided to wait until tomorrow to ask our faculty advisors for their input.

Wednesday June 26

In the morning, we continued researching the cohesin-dockerin complex and made liquid cultures of the fliC C2 domain. We also received some freebies from New England Biolabs, how wonderful. In the afternoon, we had our biweekly meeting with our faculty advisors. They have given us lots of good advice on how to solve our PCR problems and set us on the right path to amplifying the fliC C1 domain, a key threshold of completing our project. Afterwards, we worked on finding gene sequences for cohesins and dockerins.