Building better scientific thoughts
through team collaboration in synthetic biology

    To promote the progress of each other’s project, we actively communicated and collaborated with different teams, in which process we received and provided many valuable suggestions that ultimately improved each other’s project.

     This year NJTech_China aims at the improvement of extraterrestrial soils by microorganisms. They expect that after interstellar migration, the oxygen content in the alien atmosphere and soil will increase due to the influence of human activities. Therefore, it is necessary to develop an engineered bacterium that can survive and improve the soil under both anaerobic and aerobic conditions. This year, our team, CPU_Nanjing, plans to build genetically engineered bacteria that can manufacture phosphate from phosphite, thereby accelerating the evolution of phosphorus on terrestrial planets.

    After making some progress in the experiment of the project, we further exchanged each other's ideas on molecular cloning strategies. When they learned that the plasmid-overexpressed polyphosphate kinase (PPK) in our project uses ATP as a substrate, they advised us not to use high-copy number plasmids with replication origin pUC, but consider switching to low-copy number plasmids with the origin of replication pBR322 instead. They explained that ATP is required for plasmid replication and the overexpression, transcription and translation of PPK, meaning that a high-copy plasmid itself will consume a large amount of ATP [1,2]. Even if we obtain a large amount of the enzyme PPK, it will still be unable for PPK to realize the high yield of polyphosphate (polyP) due to the lack of substrate ATP. In addition, they recommended us to refer to the primary literature that initially reported this equilibrium expression strategy [3].

    Afterwards, we studied these above-mentioned literatures carefully, switched our expression vector to pBBR1MCS2, and found that the production of polyP was indeed improved. So we reported the result to NJTech_China online.

    In view of our fruitful cooperation with NJTech_China at the beginning of the project, we decided to develop Partnership with NJTech_China to promote each other's iGEM project throughout this year.

    NNU-China is an experienced team from Nanjing Normal University. Our collaboration started with an offline, in-lab meetup, where they first introduced us to their lab and the apparatuses used for their project.

    Next, we communicated about gene configuration issues involving engineered functions in each other's projects. The work of NNU-China this year is extremely challenging. They will use Yarrowia lipolytica as the chassis to introduce the metabolic pathway for the synthesis of docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) through genetic engineering, meaning that they needed to figure out every enzyme required for DHA/EPA synthesis and the genes encoding them.

    Our challenge this year is to enhance the biosynthesis of polyP, an intermediate in the phosphate manufacturing process. Because polyP consists of phosphate, more polyP means more phosphate. Our design focused on promoting the overexpression of phosphite dehydrogenase and polyphosphate kinase, and other required enzymes would be based entirely on the host's own endogenous metabolic pathways. After study and discussion, we together agreed that, regardless of the number, we were both faced with the challenge of how to combine and order the enzymes on their expression vectors. Only through sorting optimization can the optimal solution be found, thereby maximizing product yield.

    The meeting between the teams was very harmonious and pleasant. After the meeting, we exchanged souvenirs with each other.

     In September, we finished the construction of both sorts, phosphite dehydrogenase + polyphosphate kinase (phosphite dehydrogenase first) and polyphosphate kinase + phosphite dehydrogenase (polyphosphate kinase first), after which we sent the two vectors to their lab in the form of glycerol bacteria so that they can test the efficacy when needed.

    We witnessed a presentation from ShanghaiTech University at this year's Conference of China iGEMer Community online conference, and learned that they, like us, set the scene of the project on terrestrial planets.

    Since the environment of terrestrial planets may be completely different from that of Earth, the two teams will face many joint, unknown challenges. Therefore, what challenges exist and how to address them became our common topic. To that end, we have had a lengthy online discussion focusing on the elements necessary to sustain life in space. We reached absolute agreement that if we can use synthetic biology methods to convert carbon, nitrogen, oxygen, phosphorus and other indispensable elements into forms that can be utilized by life in extraterrestrial environment, it will be able to accelerate transforming terrestrial planets to habitable ones.

    Interestingly, the research contents of the two teams complement each other. This year ShanghaiTech_China is devoted to the fixation of gaseous cyclic elements carbon and nitrogen, that is, the conversion of carbon dioxide into organic matter through carbon fixation and that of nitrogen into ammonium salts through nitrogen fixation. Being similar to their plan, we CPU_Nanjing is dedicated to the oxidation of phosphorus element in sedimentary cycle. More coincidentally, we all expect to fix carbon through the photosynthesis of algae in order to produce organic matters as a carbon source for chassis.

    This productive communication highlights the importance of collaboration in iGEM competition. Both teams have had a deeper understanding of the strengths and weaknesses of each other's projects. We believe with confidence that we can supply each other with complementary products to make up for each other's disadvantages in the future, striving together to make migrations towards terrestrial planets possible.

Reference

[1] J. Vind, M.A. Sørensen, M.D. Rasmussen, S. Pedersen, Synthesis of proteins in Escherichia coli is limited by the concentration of free ribosomes: expression from reporter genes does not always reflect functional mRNA levels, Journal of Molecular Biology 231(3) (1993) 678-688.
[2] S. Birnbaum, J. Bailey, Plasmid presence changes the relative levels of many host cell proteins and ribosome components in recombinant Escherichia coli, Biotechnology and Bioengineering 37(8) (1991) 736-745.
[3] K.L. Jones, S.-W. Kim, J. Keasling, Low-copy plasmids can perform as well as or better than high-copy plasmids for metabolic engineering of bacteria, Metabolic Engineering 2(4) (2000) 328-338.