Why I did this post ? For this post I was looking for a subject, linked to Glowee, but without confidential informations, and of course enough interesting. I founded the idea when Simon the designer came for it morning coffee! So, I wanted to write a post on design, talking about it link with biology. So, in this post, I will not talk about the symbiosis of two organisms but the “symbiosis” between design and biology. In fact, both are important parts at Glowee, but also in other fields or companies. So, I wanted to talk a bit about design, and why these two fields are linked. But what is design? There is no real definition of design, because it change in function of the time, the fashion, the world or the cultures. But to describe design more particularly, the AFD (Alliance Française des Designers) give this following definition: “Design is a creative, multidisciplinary and humanistic intellectual process whose goal is to deal with and solve everyday problems, small and large, linked to economic, social and environmental issues.” And if biology needs design ? and vice versa? We don’t always pay attention to the design of objects, but for many of them, someone has thought about the shape, the size or the way to use it. It is also the case in many of instruments or machine in laboratories. Design is everywhere! In Glowee, the work of the designer is useful to create new shapes of bioluminescent stickers, new ideas of decors or new kind of utilisations of bioluminescent lights. Glowee absolutely need innovation in therm of light production but also design innovations, in order to create new kinds of products for example. Still working in the lab for biological researches, I see designers talking with the lab team, about our new researches, results and also needs for the experiments. So, the biological part is also a source of ideas for the designers.
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This blog post will be less scientific because I wanted to talk about the startup and how we work. In fact, it is quite different than in research laboratories. In this post, I will also talk a bit about our teamwork, and something important to me, which is “Do what you like to do”. TeamworkAs a startup, we have a team of designers, who imagine and create decorations and scenography. There is also the team for laboratory research, who modifies genetically the strains, tries different methods of production… And finally, there are the heads, who make decisions for the startup and for example decisions for special events. Every week we have a team meeting, where we talk about future events, one of us presents his last results, someone else presents a scientific article which can be interesting for Glowee, and often someone brings some “pains au chocolat”, essential for a good brain activity. ;) Moreover, we often share our ideas or problems about laboratory work or others things, thus we can find solutions or give hints to other team members. Of course, there are stressing moments, when we prepare an event, or just because we are a startup and we need to have good results (to be able to continue this adventure), but we work in a very good atmosphere, and everyone is part of the adventure! No typical days in GloweeFor the designers as well as for the lab team, every day is different. I will talk more about the lab because this is where I work ;). As I told you last time, I work on cloning and bioluminescence production in bioreactors. In a week I can work on both, or only one, it depends on the results, on the different needs for the events (sometimes we have to try some things before an event), but it also depends on the ideas of my supervisor and me. In fact, taking our results into account, we think about the different tests, for the bioreactor with 2 different culture media for example. I also spend time on Arduino (of course you know what it is but for those who'd need a little reminder, you'll find it here https://www.arduino.cc/ ), in order to make a light reference, with a LED and an Arduino UNO, for the experiments. We needed a light reference to analyse the pictures taken during the bioluminescence production. With this, we can see the accuracy of the camera. So, finally, every day are different from each other! Do what you like to doIt is true that “Do what you like to do” is more easy to say than to realize in certain situations or in certain countries. But I think this is essential to you, to your work, to your life and how you feel every day. I mean, wake up the morning and say “let’s go, it will be a good day” is sooo important. It is true that in certain families or in certain countries it is more difficult to access to the university you want or you're dreaming job. Unfortunately, it can be limited by money or just the diversity of schools and companies. But honestly, for those living in occidental countries we have a lot of opportunities and possibilities. So, when you found the courses, the internship, the job or whatever you like, just GO! For those who have less opportunities, if you are motivated, a lot of things are possible, never give up. Also, it is never too late to change your job or learn something new! To conclude, do what you like to do guys, it is so important. Why I did this post Still, in internship at Glowee, I really appreciate Glowee’s work, the team, what and how I learn new things. So, I wanted to share my experience and my vision of the startup work with you. Hope you completely like what you do!
The lux operon Just to remember, an operon is a DNA unit which groups several genes that operate under the signal of the same promoter. The lux operon is composed of 6 different lux genes; luxA, luxB, luxC, luxD, luxE, luxG. It was found that the genes luxA, luxB, luxC, luxD and luxE encode enzymes for light production. More particularly, luxA and B encode respectively for the alpha and beta subunits of luciferase enzyme. The function of the luxG gene is unknown for the moment. The other genes (luxC, D, E) are involved in the synthesis of the aldehyde substrate, that is to say, luciferin. The different genes and the operon organization in Vibrio fischeri are shown in figure 1. It can be noted that V. fischeri is independent, by the expression of the luciferase enzyme and the luciferin substrate. We use plasmids from an IGEM team (competition of International Genetically Engineered Machine), and it contains arabinose induction genes, allowing us to induce the expression of the lux proteins. Most of the time glucose is used as carbon source, but arabinose (also a sugar and carbon source) can also be metabolized by bacteria. So, in our project, we use glucose catabolite repression, where glucose is firstly used and when there is no more glucose in the media, cells will use arabinose. And at this point, the proteins for light will be expressed in the media. Figure 1: Scheme of the lux operon organization in Vibrio fischeri, containing luxCDABE genes. [1] My work here I started an internship in the startup Glowee since almost 2 months now. With my supervisor Antonio, I work on two main projects, the first one is about the light culture production with bioreactors and solid media, and the other one is cloning in order to improve light producing strains. Unfortunately I cannot tell you more about my work, because of confidentiality of the experiments and results. So, I will tell you about bioreactors and cloning. Bioreactors, or how to produce bacteria in big quantitiesA bioreactor, sometimes called fermentor, is a device in which you cultivate microorganisms, as bacteria, yeasts or microalgae. The final goal is the biomass and/or metabolites production. In general, bioreactors are composed of a large tank, containing cultures, and also composed of different probes and controllers. It allows the quasi-total control of growth, by measuring continually pH, temperature, aeration or optical density for example. Thereby, every data can be collected and it is possible to change the growth parameters. The bioreactor can have different volumes of cultures, 0.1 to 15 liters for laboratories, or until 1000 liters for a test before industrialization. [2. 3. 4] Figure 1: General scheme of a bioreactor. Here there is effluent tube, which is only present in continuous fed batch (see below). (https://commons.wikimedia.org/wiki/File:Bioreactor_principle.svg) There are 3 different types of culture in bioreactors;
Cloning part Cloning is a term used to describe different techniques in genetics, made to modify DNA sequences and obtain modified strains. In fact, recombinant DNA is assembled, which alter gene expression and replication. In general, DNA comes from one host strain, which contains genes for replication and from another DNA with the sequence of interest. As shown in the scheme, there are several parts to obtain cloned bacteria. The first is made to obtain genomic DNA from the bacterial strain, purify it and make a digestion with restriction enzymes. After this, you obtain fragments or inserts with the sequence of interest. In parallel, plasmids from the host are also purified and digested with the same couple of restriction enzymes. Secondly, you ligate the inserts and vectors together and transform bacteria with it. Generally, thermal shock is used to transform bacteria. Then you let the bacteria grow on medium with the antibiotic. After incubation, the colonies on the plate will be transformed bacteria because it grows in presence of antibiotics. It means that they have the plasmid containing antibiotic resistance gene, so the modified plasmid. Figure 3: Scheme of cloning, reprensenting the different steps, detailed above. See you soon ! In this post, I explained how lux operon is expressed, the different kinds of bioreactors and remind you how cloning works. In the next posts I will certainly tell you about Arduino and about my work here, but always with confidentiality rules ;) References [1] Baldwin, T., Devine, J., Heckel, R., Lin, J. and Shadel, G. (1989). The complete nucleotide sequence of the lux regulon of Vibrio fischeri and the luxAB region of Photobacterium leiognathi and the mechanism of control of bacterial bioluminescence. Journal of Bioluminescence and Chemiluminescence, 4(1), pp.326-341.
[2] Userpages.umbc.edu. (2017). The Arabinose Operon. [online] Available at: http://userpages.umbc.edu/~lrowan1/arabinoseoperon.html [Accessed 24 Mar. 2017]. [3] Griffiths, A., Gelbart, W., Miller, J. and Lewontin, R. (2017). Dual Positive And Negative Control: the Arabinose Operon. [online] Ncbi.nlm.nih.gov. Available at: https://www.ncbi.nlm.nih.gov/books/NBK21277/ [Accessed 24 Mar. 2017]. [4] Bioinfo.org.cn. (2017). Chapter 27 : Regulation of Gene Expression. [online] Available at: http://www.bioinfo.org.cn/book/biochemistry/chapt27/bio4.htm [Accessed 24 Mar. 2017]. Just to remember ;)I started my internship in the startup Glowee 3 weeks ago. The goal of this startup is the utilization of bioluminescent genes from the Vibrio fischeri strain, expressed by Escherichia coli strain, in order to bring light into streets with signaletics or showcases but also during special events.[1] During this internship, I work on the bioluminescent light, in order to ameliorate the light intensity, the time of luminescence and have the cheapest protocol. Last time I explained to you how bioluminescence works (if you didn’t read my article I invite you to read it before :) ) but also the symbiosis between the cuttlefish Euprymna scolopes and the bacteria Vibrio fischeri. Last time I teased you about the lux operon, but I will develop it in the next post. So for the moment, I will just tell you more about this amazing symbiosis. Symbiosis - the simple way to live togetherThe bioluminescent bacteria V. fischeri apparently take the control of the internal clock of Euprymna scolopes cuttlefish. In fact, the team of Margaret McFall-Ngai of the University of Wisconsin in Madison (USA) discovered that V. fischeri is implicated in the circadian cycle of the cuttlefish and not only in the light production. [2] To remind you, circadian cycles are biological cycles of 24 hours where the organism is regulated. Moreover, this rhythm regulates most of our biological and behavioral functions. Solar light plays an important role in this circadian regulation. ID card of Euprymna scolopes and Vibrio fischeriEuprymna scolopes is a cuttlefish, from the family Sepiolidae. It measures about 30 millimeters for 3 grams. [3] This organism live in the Pacific ocean, near Hawaï. E. scolopes feeds mainly on shrimps, accessible in shallow waters. Picture of Euprymna scolopes https://katiesuedavis.wordpress.com/2011/12/20/about-the-header-vibrio-fischeri-and-euprymna-scolopes/ Vibrio fischeri is a motile - gram negative - seawater bacteria. It specificity is bioluminescence, in fact this bacteria brings light into his host in the sea. The bioluminescence is allowed by the expression of proteins expressed with lux operon. Plate with bioluminescent Vibrio fischeri culture https://www.flickr.com/photos/55386616@N07/7082321451. Who, where, when . . .In our example, the cuttlefish could not shine during the night without V. fischeri. This blue light allows it to lure the predators and hunt some shrimps. [4] This cohabitation is mutually beneficial; bacteria use nutrients present in the cuttlefish and this one provide a favorable environment. But how this symbiosis works? And where does it start? V. fischeri is the only one capable of effectively colonizing the luminous organs of Euprymna scolopes. In fact, the symbiosis starts at the birth of the cuttlefish. The luminous organ of Euprymna scolopes has ciliated arms, and this traps the marine bacteria ... including V. fischeri which advances thanks to their flagellum. When the bacteria is inside the animal, the expression of several light genes is changed. For example, V. fischeri produces toxic molecules for others bacteria or in contrary attracts other Vibrios with chitobiose (major product of chitinase).[5] But Euprymna scolopes also acts on bacteria, by regulating the number of bacteria, not to produce light when it is daylight. When the night is coming, the number of bacteria increases, thanks to quorum sensing (see below). Scheme of the beginning of the symbiosis between the two organisms. Research project on this mechanismA lot of researchers tried to understand how V. fischeri acts on this bioluminescence (from a genomic and proteomic point of view). I told you about quorum sensing. But what it is? This phenomenon is induced when the bacteria releases chemical self-inducers to warn others of their presence, and when the level of autoinducers reaches a certain density the bacteria activates genes that react with proteins, here luciferase, to emit light. With this last phenomenon, researchers deduced that cuttlefish circadian cycles are regulated by the bacteria itself. Tests were carried out in the laboratory, cuttlefishes that had no Vibrio fischeri, could not become luminescent and did not produce the cycle of expression of the escry1 gene (one of the genes involved). Even by imitating bioluminescence with blue light, the cycle was not induced. In the latter case, if the bacteria is present but incapable of producing light, the cycle is activated when the dummy light is used. This proves that bacteria and light are thus essential to control the gene cycle in cuttlefish. [6] Bonus: The wonder organs of Euprymna scolopesAdded to this, Euprymna scolopes had a specific reflective organ. Composed by several parts, as reflectors, lenses or photocytes. Each part acts as a mirror, amplifying the light intensity! We keep this in mind for our project, to increase the light by this kind of systems. Scheme of a light organ in cuttlefishes, from : http://champo-tpe.e-monsite.com/pages/content/ii-type-de-bioluminescence/a-structure-des-organes.html. For the next blog posts . . .I explained from which organisms this bioluminescence comes from, and next time I will explain you the mechanism at a genetic level. For the followings posts I will talk about what I did during the internship and probably other nice stuff ;) References
Glowee’s projectFor this second semester, I started an internship in the startup Glowee! You maybe know this startup [4], which started to work with the OpenLab and the CRI. But I will tell you more about it! Everything starts in a design school in 2013 where Sandra Rey and Maelle Chassard participate to the competition Prix ArtScience. Their project was retained for it ecological and economical interest. It is their first win and the beginning of the Glowee’s story. This startup offers a 100% ecological public lighting system made from bioluminescent organisms. Which is very interesting, given that in France in 2014 the global consumption of electricity for lighting was 12%. And this number increases year after year. So there is a real need to change how we consume energy. Glowee’s team decided to create a kind of sticker, containing the bioluminescent organisms, to light showcases. For the next few years, they plan to create lightings in urban landscapes, as signage, buildings or special events. The amazing bioluminescent mechanismsSo as you understood, bioluminescence is the production and emission of cold light by living organisms (because less than 20% of the light generates heat). [2] A lot of organisms are able to produce bioluminescence, as calmars, fishes, fireflies, dinoflagellates or bacteria. In fact, there are more than 700 identified species!This phenomenon is mostly present in animals of the dark seabed, used to locate or attract their prey. Most of the light emissions belong to green-blue spectrum, giving this blue light. This wavelength can be transmitted easily through water. Pictures of dinoflagellates, an example of a bioluminescent organism, and the sea lightened by bioluminescence with it. aqueros.blogspot.com , http://blog.surf-prevention.com/2012/04/06/bioluminescence-vagues-lumiere-bleue/ But how is produced this amazing light? There is 3 different “type” of bioluminescence [6]. The first one is intracellular, generated by specialized cells of pluricellular organisms, the photocyte cells. This light is emitted towards the exterior through the skin. Also, some fish possess guanine plaques to intensify the light. The responsible organs are named photophores, composed by glandular cells, reflectors and rods that direct the light towards a single large cell that acts as a crystalline lens. The enzymatic reaction which releases the photons occurs from the glandular cells. The second kind of bioluminescence is extracellular, [2] allowed by an enzymatic reaction. The emitted light is the result of an enzyme, the luciferase, on the luciferin molecules. This catalyzed reaction needs ATP and dioxygen. The light production occurs during the oxidation phase of luciferin. You can see the following reaction equation: (Luciferin + ATP) + (Luciferase + O²) => Oxyluciferin + Photons After this synthesis, the substance is stocked in the glands of the skin or under it. After, the organism ejects it and produces light clouds. This kind of bioluminescence concern some species of crustaceans, abyssal cephalopods and fireflies. The last kind is symbiosis with bacteria, utilized during this project. This is the most common mechanism for bioluminescence. As a reminder, the symbiosis is the “Interaction between two different organisms living in close physical association, typically to the advantage of both” (according to https://en.oxforddictionaries.com/definition/symbiosis ) So, animals have small vesicles in their body, called photophores, containing bioluminescent bacteria! These two organisms live together and produce light when the number of bacteria is high. Euprymna scolopes and Vibrio fischeri, a long love storyIn this project, we studied the symbiosis [3] between Euprymna scolopes, a tiny cuttlefish from Hawaii and Vibrio fischeri a marine bacteria. This exclusive relationship allows the cuttlefish to be “invisible” for the nocturne predators. In fact, from the top you can see blue sparklings, but from the bottom, the cuttlefish becomes a furtive prey. It allows the cuttlefish to hunt shrimps and worms. Also, the unique goal of the Vibrio fischeri is growth. But without Euprymna scolopes, the bacteria could not use the nutrients inside the host and use this favorable environment for its own growth. The cuttlefish regulates the luminescence by regulating the number of bacteria, evicting them when necessary. So both organisms play a role in the survival of the other one. Picture of bioluminescent Euprymna scolopes, source; https://bionique.artbite.fr/Seiches-et-calmars-lumineux.html Glowee’s work and teasingIn this project, the team study the genes of Vibrio fischeri, the lux genes, a complex mechanism to produce light. In fact, the goal is the amelioration of the light intensity, the time of light, and the culture conditions. So, in the next post, I will tell you more about these genes and the magical symbiosis between Euprymna scolopes and Vibrio fischeri, because I only explained a part of the story [5]! If you like TED conferences and you are fascinated by bioluminescence, watch the video of the conference made by Sandra Ray, director of Glowee! https://www.youtube.com/watch?v=789IeXkOUfM More links if you are interested by this Start-up, take a look at the website! http://www.glowee.eu/ References[1] Engebrecht, J. and Silverman, M. (1984). Identification of genes and gene products necessary for bacterial bioluminescence. Proceedings of the National Academy of Sciences, 81(13), pp.4154-4158.
[2] Annual Review of Microbiology, Scripps Institution of Oceanography, La Jolla, California, (1977). BACTERIAL BIOLUMINESCENCE. [3] Mulot, R. (2017). Comment une seiche lilliputienne éclaire la recherche. [online] Sciences et Avenir. Available at: https://www.sciencesetavenir.fr/nature-environnement/comment-une-seiche-lilliputienne-eclaire-la-recherche_15466 [Accessed 20 Feb. 2017]. [4] Glowee, enlightened by the sea. (2017). [online] Glowee, enlightened by the sea. Available at: http://www.glowee.eu/ [Accessed 20 Feb. 2017]. [5] Des bactéries bioluminescentes contrôlent l’horloge interne d’un calmar. (2017). [online] Futura. Available at: http://www.futura-sciences.com/planete/actualites/zoologie-bacteries-bioluminescentes-controlent-horloge-interne-calmar-45611/ [Accessed 20 Feb. 2017]. [6] Bioluminescence - Chemistry Encyclopedia - reaction, molecule, Beetles/Fireflies, Dinoflagellates, Bacteria. (2017). [online] Chemistryexplained.com. Available at: http://www.chemistryexplained.com/Ar-Bo/Bioluminescence.html [Accessed 20 Feb. 2017]. |