Modular tools towards drop-in biofuel production by thermophiles

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Elena Martinez-Klimova, Prof. David Leak, Dr. Tom Ellis and Thomas Hargest

United Kingdom

Geobacillus thermoglucosidasius is a thermophilic bacterium that can convert monomeric and some oligomeric and polymeric components of lignocellulosic feedstocks into ethanol biofuels at high temperatures. Its application for industrial use is currently hampered by the lack of tools for rational and rapid genetic engineering. To enable design-led synthetic biology and metabolic engineering in this useful thermophile chassis we have developed a modular plasmid tool kit suitable for overlap assembly methods (Gibson) and standard restriction digest cloning. Within this kit is a genetic knockin/knockout system that utilizes green fluorescent protein as a visible marker. We are now able to use it to up-regulate precursor formation and to introduce non-native genes in order to produce the drop-in biofuel, isobutanol.