# Achievements

We have spent a fruitful –and stressful- summer working on an empirical, mathematical and comprehensive approach on the engineering pillars of SynBio. We have fulfilled the Gold Medal requirements: we have documented our standards that were submitted to the Registry of Biological Parts, and also validated them experimentally; the characterization of existing parts has been enormously improved -all the ten BioBricks used in the St2OOL project are combinations of existing parts of the registry, and we believe have performed one of the most complete characterizations of existing Registry parts in terms of stability, standardization and orthogonality; and, last but not least, we have developed an original Human Practices approach dealing with Intellectual Property in Synthetic Biology in the context of Responsible Research and Innovation.

## Yes, we could!

We have performed an unprecedented research effort to determine in detail how biological parts behave under different conditions. We have used several hundreds of Petri dishes and fluorescence –among other- measurements, which have allowed us to finely characterize the behavior of ten selected SynBio devices. Our main results are the following:

1. Escherichia coli is, as expected, an extremely strong chassis for SynBio, as its is able to survive in a very wide rage of environmental stresses, of which we have tested pH, salt, Mars-like vaccum, temperature and radiation. The only factor to which E. coli proved very sensitive to was UV light. By contrast, Biobricks behavior was dependent on environmental conditions: sub-optimal temperatures and vacuum dramatically decreased Biobrick output.

2. Emergent properties. Even in 2014, many new species are discovered hidden in the jungles of our Planet. Similarly, we have been lucky to report here an unexpected emergent property in the best-known bacterium on Earth: thermal oscillations resulted in a significant increase of E. coli DH5-alfa growth . We are still finding out how. Surprisingly, fatigue of materials in biology is not always a deleterious process.

3. Standards not that standard. We were shocked to see only Biobrick 5 behave in a relatively standard way when its output was compared within six different E. coli strains. Variation in the output was the norm, not the exception in what we expected to be standard parts.

4. Orthogonality bench testing. We used a range of techniques (fluorimetry, cell citometry, protein electrophoresis and proteomics) to find out whether one of the simplest device combination possible (two different reporter proteins under the control of the same promoter and on the same plasmid) behave in a orthogonal way. The hypothesis was that, if so, the relative outputs should be approximately equal. Again, we found that engineering assumptions could not be validated: co-transformants with two Biobricks displayed an asymmetric behavior indicating a clear lack of orthogonality, and a huge cell-to-cell variation in output signal was measured .

5. What is the effect of a genetic modification in the proteome? We performed a proteomics analysis of one of our Biobricks (Bb1, GFP) and compared it with a wild-type strain and an empty plasmid-bearing one. Three replicates were analysed. The results reveal a small impact in the proteome profile of both the empty plasmid and the plasmid with Biobrick1. Only chaperones and other few proteins were affected. To the best of our knowledge, this is the first proteomic demonstration of the effect on the proteomic architecture of the introduction of a simple SynBio device.

6. Bridging gaps in IP. We wanted to contribute to the IP controversy in a new way. We propose a marriage between the concepts of Responsible Research and Innovation and the Diverse Ecology scenario, being the former a key tool to choose within the multiple ecology of IP forms. We have contributed to that goal by creating dictionaries of SynBio terms for lawyers, and of legal terms for Synthetic Biologists, and by making a politically incorrect proposal: why not using Maths to help experts on IP take their decisions?

7. Modeling and wetlab, together. We modeled the four legs of the St$^2$OOL: from the stability of Biobricks (which proved to fit the shape of our actual data) to the Human Practices equation. We studied in depth orthogonality and the behavior of plasmids co-transformed into a host cell; we have created a software to generate promoter sequences with a desired level of expression and compute the stability & standardization (St$^2$) index and proved able to both model and forecast a range of biological experimental data.

8. On weakness and strength. Our experimental results and modeling analysis confirm that engineering pillars cannot be taken for granted in SynBio. A new bioprospection point of view is thus needed. We have developed a DNA extraction robot “The Tool” to help us carry out a functional metagenomic approach directed at identifying natural DNA parts with particularly standard and stable behaviors. If engineering assumptions are not met by the Biobricks we tested, let’s isolate/build new ones with the desired properties.

## Beyond scientific achievements

We really had fun! In our “Art & More section” you’ll find Paula’s great watercolors for the wiki, the poster and the presentation; Kristie’s epigenetic music interpreted as what she calls “genomic serialism”; and videos such as the YMCA parody “The St2OOL is going to i.G.E.M.”

## And finally, the St2OOL quote

"I never use a piano stool. I always use a drum stool. Because I feel that when you're down there, you're playing in that way you're supposed to. I like to be above it." Jamie Cullum