Improving the performance of solventogenic clostridia using CRISPR cas

Project

Improving the performance of solventogenic clostridia using CRISPR cas

The discovery of the CRISPR cas technology opens new doors for industrial biotechnology. Using CRISPR cas based tools, we aim to increase the solvent production of solventogenic clostridia by disrupting the sporulation.

Context

The growing global population and the emergence of developing countries drives the demand for consumer goods. However the manufacture of these products is entirely resting on the petrochemical industry. Since fossil fuel resources are limited, there is a great need for sustainable alternatives. Acetone-Butanol-Ethanol (ABE) fermentation is one of the bioprocesses explored by industry and academics. ABE fermentation enables the conversion of a wide range of carbon sources such as CO2, organic waste and complex sugars to solvents. This conversion is executed by natural solvent producing bacteria, called solventogenic clostridia. They are gram positive anaerobic bacteria that form spores. They are characterised by their ability to produce organic solvents such as acetone, ethanol or butanol. These chemicals are commonly sold as paint thinners, glue additives or for perfumes. However, the ABE fermentation process is currently not cost-competitive with the petrochemical process used. Thus, researchers are looking for ways to improve the strains used. One of the approach investigated is the engineering of a non-sporulating strain.

Sporulation cell differentiation process triggered when the cells are in an unfavourable environment (presence of oxygen, high pH variation). Spores protect the genetic material and ensure the preservation of the specie until favourable environmental conditions arise. However sporulation is seen as a hurdle, in industry, for several reasons: it occurs during the fermentation, stops the solvent production and is energy costly for the cells. Moreover, researchers believe that preventing the cell from sporulating could increase the efficiency of process by redirecting the energy used for sporulation to the solvent production.

Aim

Our goal is to use and adapt CRISPR cas editing techniques to generate non sporulating strains of C. beijerinckii and to characterise them.

Techniques

Depending on the project the student will have the opportunity to learn several techniques such as:

  • Culturing of anaerobic bacteria
  • Transformation of Clostridium strains
  • Construction of vectors and cloning in E.coli
  • Use of CRISPR-Cas based genome editing in Clostridium
  • Characterisation of mutants ( microscopy, fermentation, RTqPCR)

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