thanks! I guess a good compromise would be 500ml for now.

Sure, I will collect the links for the plastic components from Aliexpress and update your sheet soon.

Largest would be 500ml without changing much. Maybe will add three or four heating-pads, thus adding another two motor drivers. Plus, scale up the size of the heating jacket model. then 3D print it.

If you want go higher (1-5L) then you may use the old heating system, through MOS and step-up voltage circuits, with two larger heating pads. Also will need to add the temp sensor inside the culture bottle not outside. you will ignore the 3D printed heating jacket (replaced with larger heating pads).

Only the stirring parts (3D printed base) will need to resized, and the stirring fan, also need to buy a bigger size.

Beyond 5L ---> (<10L), you will need to get bigger pumps (Stepper motor 12V), Glass container for the culture need to be carefully fabricated.

thank you for the comments in the spreadsheet. could you please take a look at the 'fluidic connections' sheet as well (the second sheet in the same document). I really couldn't find those components :( re volume: initially I'm aiming for a small container size and to verify that indeed a culture can be sustained in the reactor. then, I'd like to design a lighting unit with a much larger container but using the same principle. but as a first step, a smaller one is good enough. having said that, what would be the largest container that would 'just work' with the system you designed, without changing too much?

You are welcome :)

Yes, any custom glass shop can fabricate the culture bottle and the OD tube, the PDF file has the detailed sizes.

You planing same culture volume as in the design ? (~20 to 50 ml) ??

Check the google sheet u sent me, i commented there some parts.

SSRA tagging - sounds quite interesting. at the same time, it would also be interesting to have a similar optogenetic approach to luminescent organisms

Thank you very much for your help on the reactor components. I guess 2 reactors would be fine, so that one can run A/B tests in parallel. Thank you for the 3D design for the water container and the Alibaba / Aliexpress links. Re the OD tube - how is that made? In a custom glass shop?

Yes, you are right, but SSRA tagging will take care the expressed protein, by inducing the the proteases activity, and you can control how long the expressed protein will be there by optimizing the SSRA tagging on the C-terminus of your protein. This way you can control the duration of specific protein occurrence inside the cell.

Sure, pleasure to help, but let me ask you first, how many reactors (cultures) you want ?

If you just looking for bioreactor with one culture, then there are a lot of details and parts you can just ignore it.

Also the water container for the heater, you can now replace it with 3D printed part, I can send you the design.

The OD tube, is made of glass. (any type is okay, Pyrex is fine).

I will send you Alibaba or Aliexpress links for the items you need. I wait your answer about how many cultures you require, and i will prepare list for you with links from two suppliers.

re the optogenics / GFP: yes, you can turn off the gene, but the GFP (the protein itself) is already there and will be there for quite a long time (even after the bacteria dies). in that sense, the 'pixel' is not turned 'off', as it will shine back green through the GFP that's there. turning off the gene does not remove the protein that is already there

I started to put together a BOM / purchase list based on your PDF, that I can execute from Switzerland, where I am based. See here: https://docs.google.com/spreadsheets/d/1wLDycBLz__aUSo0zqZNQPtR7kvVLh2ALJKqYuE9Uwrs/edit?usp=sharing

I'd like to ask for your help in this - some parts I could not identify, for some parts I found 'similar' items, and some parts I found the exact same. most of the amazon links for the electronics show 'currently unavailable' products :( . for the fluidic connectors, I coudln't find most of them. unfortunately I don't know Korean, so I can't really navigate Korean sites. I also wonder what else should one buy in addition to what I listed? The Main Box / Water Box - is that something one builds from acrilic boards? The culture bottle - how is that built? How about the OD tube?

I see. Have you thought about the energy consumption for such application, I just guess in that case u may consume more power (for aeration) than what you actually produce as a light.

There is a German group i read before they tried almost similar system with Micro-algae. I don't remember their name, but they hit some news paper at that time.

However, who knows, usually the real deal important applications for any idea appear during the development.

For the optogenetics, actually you can turn on/off (any gene of interest including GFP or any other) using the operon systems, plus also C-terminus protein tagging SSRA would adjust the timing and duration for the illumination.

thank you

my target with PP is to create a sustainable light source based on a living organism. PP itself is luminescent, and will produced light when it's in a growth phase and is oxygenated. basically light production can be turned on or off through oxygenation. I've made this happen with 'manually grown' liquid cultures, but never got to a stage to have a 'sustained' culture. the end goal is to create an aesthetic installation that is sustainable and can produce light.

of course in the long run the best would be to increase light quantity, and even better is this could be done with a plant or similar, instead of a liquid culture. but that's way beyond of my current means. re the above optogenetic example: looks interesting. OTOH, inducing GFP production based on optogenetics seems to be a 'one way street' - the GFP will remain there for a long time, e.g. one an turn it 'on' but cannot really turn it 'off'. now, if we could induce luminescence with similar means, that would be interesting...

Sorry, i notice that website is urs, okay, i got your point about bio-displays, very interesting idea actually,

Would you take a look on this photo and publication below:

This been published in https://www.nature.com/articles/s41589-020-0639-1

Its actually based on bacterial optogenetics system. its more programmable, and u can create a lot of interesting stuff with it.

Let me know what is your final target of experimenting with PP bacterium, we may cooperate to some extend, I am interested in DIYs and maker stuff. :)

Sure you can increase the RPM of the aeration pump,

Scaling up to what volume ?

you can reach up to 5 liters with the same peristaltic pumps (just need to change the air pump to stronger one). Otherwise u may need shift to stepper motors for the peristaltic pumps with same motor drive. And also consider bigger air pumps.

Sounds excellent, thank you. I have two additional questions:

Photobacterium Phosphoreum needs to be heavily oxygenated to produce significant amounts of light. See for example here: http://biodisplay.tyrell.hu/2015/08/04/on-the-way-to-a-light-source/ . Would you have suggestions on 'higher level' airflow into the container?

Would you have suggestions on how to 'scale up' the incubator in size?

Yes, almost that way, you can expect some fluctuation in the OD readings due to the dilution of each waste-feed cycle,

However if you looking for the steady state, you can tight the OD-based system, by modifying two lines of the code (I can send/guide u through the sketch if u need my help on this), by this modification u can achieve the steady state, with the lowest OD fluctuation.

thank you thus how this works os that one reads the optical desnity (OD) sensor with the arduino in the reactor, and 'identify' the exponential / deceleration / saturation phases, and then apply a waste / feed cycle based on that

that is, release waste and supply feed to get at the lower end of the exponential cycle (about -2 on the above chart), then 'wait' until about the higher end of the exponential phase (about -0.5 on the above chart) - reaching that, waste is released & feed supplied to get back to the lower end (about -2)?

Hello Akos

Thanks for your words, actually i did not test the design with PP bacterium, i normally use with E,coli, or other non-fluorescence strains. However, I guess it will be no problem to cultivate it in that design. You may consider adding another photo-diode for the measurements of luciferin light emitted by the PP.

The OD (turbidity) sensor in this design is IR-based which may conflict with the auto-luminescent of the PP.

This photo-diode may work fine in your case i guess (https://www.amazon.com/PHOTODIODE-Hermetic-Enhanced-Photodiodes-Photoconductive/dp/B07NJ6P18Z).

For the second question, "its peak" as you mentioned means log (exponential) phase of the bacterial growth curve. you can first characterize the bacterial strain in shake flask for 24 hr and measure the OD every fixed time interval thus you can keep tracking the bacterial growth curve pattern, it should be something like this :

Once you get that curve in your hand, then you can adjust the reactor incubation period (Feed/waste cycles) to Log (exp) phase timing.

Other than that, the software of the design also have the OD based feed-waste cycle, which you can specify one OD once the sensor reach it, it will automatic way start the feed-waste cycle.

BTW1, the Feed-waste cycle in this design is ordered as (Waste --> Feed) to govern the culture volume.

BTW2: you can characterize the bacterial strain in that reactor design (Get the Growth curve pattern), I usually do this while starting any new experiment, even with same strain.