Science Discovery Day

National Science and Engineering Week exploded in Fife with a regional "Science Discovery Day". Team St Andrews worked to convey fundamental concepts in Genetic Engineering and Synthetic Biology in new and exciting ways. The interactive "Codon Game", 3 - Dimensional Visualizations of DNA and DNA polymerase and the display "E. Coli: Under the Microscope" were well received. Children and adults alike were fascinated when DNA was extracted from bananas using everyday kitchen equipment.

June

Week 1

4 June 2012

Full time work on Team St Andrews' iGEM Project finally begins! After an initial Group Meeting, two thirds of our student members continue in depth research into those project ideas generated previously; the rest begin work on Team St Andrews' Wiki.

5 June 2012

"CLC bio" and "Epoch Life Science" are the latest businesses to offer support to Team St Andrews. The former promises CLC bio Main Workbenches to members of the team while the latter offers products and expertise in DNA/ RNA preparation for molecular manipulation.

Team "ω-3" weekly summary

Research into polyunsaturated fatty acids has been active for over 15 years, with successes varying from creating transgenic plants enriched in ω-3 fatty acids to expressing an entire synthetic pathway from a gene clusters extracted from marine bacteria. We want to expand on this area of research by attempting to express a aerobic synthesis of unsaturated fatty acids in E.coli, which has never been done before. By combining the genes of the cyanobacteria Synechococcus and trypanosome T. brucei into E.coli, this vector should be able to synethise a unsaturated fatty acid up to at least 22-carbon length!

This week, our team has been focusing on preliminary research by reading relevant scientific papers and understanding the various pathways and methods of recombination. We’re focusing on groundwork research done in the early 1990s that appears to have fallen off the radar, and are excited to see where this path will take us!

Team "Metal Binding Protein" weekly summary

"After a short meeting (which was dominated by the excitement of receiving our very own iGEM pins) we decided to split our iGEM family of nine into three groups. Josi, Veronica and Yiwang were to look into Omega-3 production; Constantine, Michael and I were to investigate Metal Binding Proteins and Antti, Alexey and Zoe were to focus on setting up the wiki.

The wiki team duly produced a sophisticated template.

The research teams, however, were unanimously agreed that biology is hard. Our motif quickly became standardised. Read, Think, Re-read, Re-think, Repeat.

After three days, our previously nomadic team settled in the University’s new Biomedical Sciences Research Complex. This proved to be a good move as it was followed by breakthroughs on all fronts. Metal binding peptides were located in their tens and the decision was made to try and express them on the surface of cells in display proteins as well as cytosolically. A membrane protein was found, preliminary peptides selected, and a primer design tutorial set up for the following Monday. All this success left us with one very important and so far unanswered question- how exactly are we going to assay all this?" (Team Member Hannah Taylor, Week One Report, 12/06/2012)

Week 2

12 June 2012: Team "Metal binding protein"

· Using E. Coli DH5-alpha: good at replicating vectors but not good at expression · Using pGEX-6P-1 vector with internal GST tag, selected with ampicillin http://ecoliwiki.net/colipedia/index.php/pGEX-6P-1 (further information via the “source” link) · Purpose: o First day: incorporate desired vectors into vector-replicating E.Coli and incubate them on selective plates to select out recombinant colonies with antibiotics o Second day: incubate the selected colonies in large quantities for vector harvesting o Third day: harvest vector from cell pallet thus pure vector miniprep.

12-15 June 2012: Team "Metal binding protein"

Using E. Coli DH5-alpha: good at replicating vectors but not good at expression · Using PET-15b vector: with internal His tag, selected with ampicillin http://ecoliwiki.net/colipedia/index.php/pET-15b (further information via the “source” link) · Purpose: o First day: incorporate desired vectors into vector-replicating E.Coli and incubate them on selective plates to select out recombinant colonies with antibiotics o Second day: incubate the selected colonies in large quantities for vector harvesting o Third day: harvest vector from cell pallet thus pure vector miniprep.

Omega Squad Week 2 Summary

This week, we took the first steps to put our grand scheme into laboratorial action! It also occurred to us that if we started calling ourselves DJ Omega 3 & the Fatty Acids, we would intimidate the other squad.

First of all, we chose a vector to work in, pET-15B. We started looking into promoters suitable for both the vector and the chain of genes we want to express, and figured out how to use the Registry of Standard Parts. The next step was to design primers for all four of the genes of our pathway - and that was certainly a steep learning curve!

Preliminary tasks done, let’s head to the lab! We carried out a transformation, growing the vector in a DH5-α strain of E. coli with ampicillin as the antibiotic marker. Well, we attempted to carry out a transformation. It failed, much to the glee of the Metal Mickies. Fortunately, we were able to use some pET-15b grown in E. coli for a different purpose. The colonies were allowed to grow, and we then carried out a mini-prep to isolate the vectors. Visualization was done by running the DNA on an agarose gel and using UV light. Once we were sure the transformation was successful, it was time to add the restriction enzymes!

Also, a PCR was run with the genomic DNA of Leishmania major, the trypanosomatids that are providing us with the 4th gene of our synthesis pathway. We ran 5 samples total: 3 using high-fidelity PCR at different temperatures, and 2 of Clontech’s PCR kits at 2 temperatures. We added the primers for Δ6 elongase (GenBank LmjF32.1160). This gene is around 1.200 kb large. The results show that the Clontech kit run at 48° C gave the strongest result.