From 2014.igem.org

School Year

Notebook | June | July | August

	Weeks Thirteen and Fourteen- September 1-14 Sept 1-7 BioBricking officially starts. All protocols established, including Double and Multiple Digestion of DNA, Fast Digestion of DNA, and Reaction Set-up for Digestion of Multiple DNA Samples. Our specified Restriction Enzyme Digest Protocol is created based on the protocols listed above. Ben sequence verified the new modified plasmids and tested them. They did not fluoresce so designed new primers for swapping out the promoter, which was the original suspect. He started cloning later in the week and sent in for sequence verification. Sept 8-14 Caroline continues on Biobricking. Richard continues the acetylene reduction assay to test nitrogenase activity of JM109 and WM1788 under known optimal conditions. Ben gets back the sequences for swapping the Ptet promoter. Only the positive control was successfully changed, as some of them had a few deletions and mispriming. He ran an experiment to test for the fluorescence of the new pTet and found that it fluoresced. Ben also started design of primers for biobricking the light repression mechanism, and the newly functional positive control.
	Weeks Fifteen and Sixteen- September 15-28 Sept 15-21 Caroline continues on Biobricking. Richard finishes the acetylene reduction assay for nitrogenase activity and starts data analysis. Ben started biobrick cloning using the new primers that came in. Did a double digest with the biobrick restriction enzymes to assemble with a part that we had received from the registry. The cloning attempts did not work at all. Sept 22-28 Caroline continues on Biobricking. Richard finishes acetylene reduction assay data analysis. The team started organizing content to place on poster for the Undergraduate Research Symposium at Washington University. Ben and Cheryl started the biobrick cloning project over again by retrying different concentrations for ligation. We tested the biobricked plasmids using new sequencing primers and saw the correct prefix but no suffix. We ordered new primers to start swapping the entire reporter gene expression from the positive control to our light plasmids. Prepped the swapped pTet for sequencing.
	Weeks Seventeen and Eighteen- September 29th- October 12th Sept 29-Oct 5 Caroline is unable to continue the biobricking process. Constructs contained illegal restriction enzyme sites and thus will not be allowed to be sent in to the registry. The team worked on the poster of our iGEM summer project for Washington University’s Undergraduate Research Symposium. Ben started cloning for biobricking over again with a new protocol CPEC based on Andrew’s protocol. We did these in conjunction with modified versions of ligation and found the correct sequencing pcr bands with only the CPEC versions. Discarded the positive control biobricking. Sent in the hybrid and normal promoter for sequencing. Oct 6-12 Poster for Undergraduate Research Symposium at Washington University is finished and printed on Oct 7th. Ben sequence verified his biobrick products and shipped the product to iGEM registry on Oct 8th, 2014. It arrived at iGEM registry on the 9th. Ben also ran two experiments: the first to test the newly cloned reporter protein mechanism, which worked finally, and the second to test the light induction. Currently analyzing the data to figure out how to best present the information. Poster presentation for Undergraduate Research Symposium is on Oct 11th , 2014.

Team:WashU StLouis/Notebook/September

From 2014.igem.org

School Year

Notebook | June | July | August

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