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Optimization of Xylose Fermentation

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Optimization of Xylose Fermentation

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    1. Optimization of Xylose Fermentation NISHANT KUMAR M.Sc ii Roll no 06530006 NISHANT KUMAR M.Sc ii Roll no 06530006

    2. INTRODUCTION Xylose or wood sugar, one of the most abundant carbohydrates in nature, is an aldopentose a monosaccharide containing five carbon atoms and including an aldehyde functional group.

    3. XYLOSE METABOLISM

    4. Xylose Metabolism & S. cerevisiae Native strains of S. cerevisiae do not assimilate xylose. In xylose fermenting yeasts, xylose is reduced to xylitol by NADPH-linked xylose reductase (XR) and xylitol is oxidized to xylulose by NAD-linked xylitol dehydrogenase (XDH). Subsequently xylulose is phosphorylated by xylulokinase (XK) for assimilation via the pentose phosphate pathway.

    5. Xylose Metabolism & S. cerevisiae The discovery that S. cerevisiae can ferment xylulose initiated metabolic engineering of xylose fermentation in S. cerevisiae. Heterologous expression of genes (XYL1, XYL2, and XYL3) coding for XR, XDH, and XK from P. stipitis enabled S. cerevisiae to utilize xylose as a sole carbon source.

    6. Shuffling of Promoters for Multiple Genes To Optimize Xylose Fermentation in an Engineered Saccharomyces cerevisiae Strain Chenfeng Lu and Thomas Jeffries APPLIED AND ENVIRONMENTAL MICROBIOLOGY, Oct. 2007, p. 60726077

    7. Multiple-gene-promoter shuffling (MGPS)

    8. Fuse selected promotor to target genes

    9. Concatenate each promoter-gene product with different combinations

    10. Screen for phenotype of interest among these transformant

    11. AIM Optimization of xylose fermentation by shuffling the promoters for GND2 and HXK2 with the genes for transaldolase (TAL1), transketolase (TKL1), and pyruvate kinase (PYK1) in the Saccharomyces cerevisiae strain FPL-YSX3.

    12. Selection of promoters The promoters for this study were chosen on the basis of the native gene expression that they control & promotor strength was validated by using lacZ expression & -Gal assay.

    13. Correlation between expression array data and -Gal activities resulting from promoter strengths of selected genes.

    15. RESULTS

    16. RESULTS.

    17. CONCLUSION The optimal expression levels for TAL1,TKL1, and PYK1 were identified by analysis of volumetric ethanol production by transformed cells & the optimal combination for ethanol production was found to be GND2-TAL1-HXK2-TKL1-HXK2-PYK1

    18. THANX!!!!

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