Chloroplasts hold a massive potential as an alternative protein expression system, due to their outstanding expression yields, diversity of post-translational modifications and auto/mixotrophic lifestyles of plants and microalgae.

Various proteins have already been successfully expressed in chloroplasts, including:

• monoclonal antibodies
• antigens
• anti-toxins
• growth factors

Transgene expression in microalgae can total up to 30-50% of a cell’s dry biomass, as unlike in higher plants and mammals, metabolic energy in microalgae is not directed towards maintaining complex differentiated structures.

Research in C. reinhardtii chloroplasts can help achieve the following aims:

• increase yields of oils for biofuels
• elucidate photosynthetic machinery
• improve C fixation to combat the international food crisis

Due to extensive evolutionary conservation of the chloroplast genome, research in the chloroplasts of microalgae, such as Chlamydomonas reinhardtii, is likely to be applicable to studies of other plants. On the other hand, Chlamydomonas reinhardtii is much easier to grow and maintain than higher plants, such as Marchantia polymorpha.

Chloroplasts engineering is currently underexplored due to:

• Time issues — it takes 2-3 months to achieve homoplasmy, as state where all chloroplast genome copies have been transformed and when experimental results can be obtained
• Experimental cost — chloroplasts can be reliably transformed almost exclusively by biolistic devices, and commercial biolistic devices are very expensive.
• Lack of modular chloroplast genetic parts available — research is more time-consuming and cumbersome.

Our project aims to tackle most bottlenecks and democratise algal biotechnology:

• Library of chloroplast parts for C. reinhardtii — to facilitate the assembly of synthetic constructs using Phytobricks standard
• Open-source biolistics device — for cheaper chloroplasts transformations
• Open source microalgae growth facility — to growth algae in an affordable way
• Novel Cas9 strategy — to accelerate the process of achieving homoplasmy
• Modelling of Cas9 dynamics — to predict the time our Cas9 stategy would take to transform all copies of a chloroplast’s genome