Expression cartridge for the transformation of eukaryotic cells, method for transforming eukaryotic cells, genetically modified organism, method for producing biofuels and/or biochemicals, and thus produced biofuel

Abstract

The present invention describes the expression cassette for transforming eukaryotic cell which comprises the peptide encoding non-natural sequence of nucleotides with xylose isomerase feature (SEQ ID NO: 1), optionally also comprising other genes of pentose phosphate route. Additionally, it is described the microorganism filed under the number DSM28739, which, in addition to the above-mentioned modifications, also present genetic modifications from adaptive evolution. The described microorganism shows efficient consumption of xylose and conversion of ethanol when compared to its correspondent without said genetic modifications and mutations from evolution. It is also described the process for producing biofuels e biochemicals, preferably ethanol, mainly from the lignocellulosic portion of the vegetal biomass. Biofuels, preferably ethanol, and biochemicals produced by the process of the invention are also described.

The invention claimed is: 
     
       1. A genetically modified microorganism, wherein the genetically modified organism is Saccharomyces cerevisiae DSM28739. 
     
     
       2. A method of genetically modifying a yeast belonging to a yeast genus, said method comprising introducing a sequence of nucleotides encoding a peptide with xylose isomerase function, into said yeast; using homologous recombination, selectively replacing codons present in said yeast with codons present in Orpinomyces, to achieve a Codon Adaptation Index of at least 0.79; using evolutionary engineering in order to generate random mutations which favor xylose conversion to biochemicals and/or biofuels; and recovering a genetically modified yeast.  
     
     
       3. The method of  claim 2 , wherein said Codon Adaptation Index is from 0.79 to 0.91.  
     
     
       4. The method of  claim 2 , said method further comprising a step of fermenting a sugar, utilizing said genetically modified yeast, to generate a fermentation product.  
     
     
       5. The method of  claim 4 , wherein said sugar is xylose.  
     
     
       6. The method of  claim 4 , wherein said fermentation product is ethanol.  
     
     
       7. The method of  claim 4 , wherein said sugar is xylose, wherein said fermentation product is ethanol, and wherein yield of said ethanol is at least 0.45 grams ethanol per gram xylose consumed by said genetically modified yeast in synthetic medium comprising said xylose.  
     
     
       8. The method of  claim 4 , wherein said sugar is xylose, wherein said fermentation product is ethanol, and wherein volumetric productivity of said ethanol is at least 0.67 grams of ethanol produced per liter per hour in synthetic medium comprising said xylose.  
     
     
       9. The method of  claim 2 , wherein said yeast genus is selected from the group consisting of Saccharomyces, Scheffersomyces, Spathaspora, Pichia, Candida, Kluyveromyces, Schizosaccharomyces, Brettanomyces, Hansenula, and Yarrowia.  
     
     
       10. The method of  claim 9 , wherein said yeast genus is Saccharomyces.  
     
     
       11. The method of  claim 10 , wherein said genetically modified yeast is Saccharomyces cerevisiae DSM28739.