Project Timeline

January

• 2013-01-15 Budget completed.
• 2013-01-29 Human Practices: Workshop for highschool students.

February

• 2013-02-25 Project selected: Conversion of ammonia to nitrous oxide in E. coli.

March

• 2013-03-11 Biobrick workshop hosted for KU and SDU.
• 2013-03-13 Project kickoff meeting with supervisors.

April

• 2013-04-15 Team contract written and signed.
• 2013-04-28 Human Practices: Held second workshop for highschool students.

May

• 2013-05-29 Hello World pilot project kicked off.

June

• 2013-06-04 Team competency mapping.
• 2013-06-09 Work on wiki started.
• 2013-06-29 Team BBQ.

July

Week 27

• Hello World project complete and successful. From a cytoplasmic extraction, we have observed GFP in the periplasm and RFP in the cytoplasm.
• Conducted the toxicity experiment and completed data analysis: average ammonia concentrations in wastewater are not toxic to E. coli.

Week 28

• Designed, ordered and received primers to extract genes from Nitrosomonas and Pseudomonas.
• Began work on cloning genes out of Nitrosomonas europaea and Pseudomonas aeruginosa. All isolations successful so far except for Nir.
• Held the first hackaton to work on the wiki.

Week 29

• Performed first experiment to investigate the anaerobic stability of nitrite in native E. coli.

Week 30

• Ordered new primers to extract Nir gene in two pieces.
  
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• Despite successful PCR and transformation of AMO, HAO and cytochromes, we only have viable colonies for the cytochromes. Expect that this is due to the fact that we are expressing periplasmic and membrane proteins, which is causing excessive stress on the cells.
• Began work on pBAD synthetic promoter library so that we will be able to induce a low level of expression of our periplasmic genes.

Week 31

• The week no PCR worked -- ran 96 PCRs with varying conditions to debug what is not working.
• Saturday BBQ with KU.

August

Week 32

• Conducted the experiment to characterize anaerobic production of N₂O in native E. coli.
• Began work on kinetic models of the aerobic and anaerobic pathways.
• Began putting together our presentation.

Week 33

• Began having daily meetings.
• Transformed arabinose inducible promoter for use with pBAD synthetic promoter library.
• Amplified one Nir fragment with USER primers (changed DMSO, Mg, temperature, etc), one left to go.
• Repeated the experiment to characterize anaerobic production of N₂O to try to reproduce results.
• Held the second hackathon.

Week 34

• Ran first biolector experiment to characterize pBAD synthetic promoter library.
• Completed first draft of presentation.
• Completed first draft of poster.

Week 35

• Attended Scandinavian meetup in Odense hosted by SDU team. Thank you SDU for a great weekend!
• Conducted another set of biolector experiments to characterize more colonies from the pBAD synthetic promoter library.
• Sent pBAD SPL samples for sequencing.
• Conducted experiment to characterize AMO and HAO transformants. Found that the AMO transformant is using ammonia faster than a nontransformed E. coli control!
  
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• Decided on project title: Requiem for a Stream.
• Wrote the project abstract.
• Reviewed presentation with Julie, who gave us very good advice on pitching our project.
• Completed safety form.

September

Week 36

• Made biobricks for AMO and cytochromes (are going to sequence these), the others are pending.
• Completed biolector experiments for the pBAD synthetic promoter library. We have constructed promoters that when induced, show higher activity than the constitutive promoter, and when not induced show very low activity.
• Began constructing a page to describe the [http://parts.igem.org/PBAD_SPL pBAD SPL in the parts registry].
• Began modelling the design of a reactor using a continuous stirred tank reactor.
• Began researching physical reactor design.

Week 37

• Verified biobricks with colony pcr using the sequencing primers.
• Human Practices: Attended BioPeople event "The Basics and the Best" on IP and patent law to discuss the impact of the US Supreme Court ruling on Myriad Genetics on synthetic biology.

Week 38

• Filmed "Bricks of Knowledge" video on USER cloning for KU team.
• Mailed Biobricks! (tat signal peptide, AMO, cytochromes).
• Conducted Bioreactor experiment.

Week 39

• Held hackathon to complete wiki.
• Conducted experiment to characterize Nir transformant.

October

Week 40

• Safety forms were approved on September 29, 2013 by the iGEM Safety Committee.
• Completed wiki.