Hi Fede,

Thank you :).

So to answer your questions 

We are planning to do intracellular expression strictly within the chloroplast. We are actually doing this to avoid many of the issues that occur because of nuclear expression, and it actually helps reduce the layers of complexity. I'll explain why.

There are a lot of advantages to expressing genes in the chloroplast because of the bacterial-derived expression machinery and the membrane bound structure of the chloroplast. 

1. Since the chloroplast is inherited maternally, there is a low chance of a foreign gene escaping in nature,

2. Since there are multiple chloroplast copies in the cell, there is a higher level of expression

3. Unlike expression from the nucleus, gene silencing doesn't seem to happen in the chloroplast

4. The chloroplast has enzymes that are capable of facilitating disulfide bond formation in proteins. This allows a wider variety of proteins to be expressed stably. 

5. Foreign proteins that would normally be toxic to the cell can be encapsulated within the chloroplast membrane

6. Any bacterial genome can be inserted into a chloroplast genome since it is of bacterial origin itself.

7. The cell wall and chloroplast membrane help facilitate delivery of oral vaccines. The cell wall allows a protein to survive the digestive track and allows the protein to be absorbed through the stomach. This is ideal for oral vaccines.

I encourage you to study the scientific literature here, here, and here if you want to know more.

Our reason for using insulin is because there is a lot of literature to guide the experiment, so we know it's been done before. Also, if oral delivery of insulin ever becomes possible, algae might be a good way to do it. you never know.

I actually have not considered cyanobacteria, but given that they are very similar to chloroplast, they would probably be great test organisms as well. You brought up something that might be really helpful that I had not thought of. Thank you :). 

I hope this has answered your questions. Please let me know if there is anything else you'd like to know.

Antonio

Hi Antonio,

Cool project.Nice to see somebody else working with algae, especially given the topic.

I may have some extra dough, even though is cyanobacteria based (metabolic engineering and synthethic biology)

I have some questions for you, hope you can answer :)

1) do you plan intracellular expression or extrusion into medium of insulin? 

2) are microalgae suitable (i.e. genetic toolbox for protein expression and/or extrusion)? (plastidial expression may add layers of complexity to your system as well)

have you considered cyanobacteria for the study?

Admirable project, wish you all the best!

Cheers,

Fede

Il giorno lunedì 7 dicembre 2015 00:31:33 UTC+1, antomicblitz ha scritto:

Hey everyone!

I wanted to take this opportunity to share a cool project that I think could be really exciting and I want to get the community involved.

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