Two genes XL1 and XL2 encoding for pentose utilization were isolated from the genomic DNA of P. stipitis (CBS 6054) using Bioinformatic tools and 2 primers each were designed to span the full coding region of these genes with attached restriction sites. PCR reaction was carried out on 120ng of the isolated genomic DNA for 30 cycles using the DNA taq polymerase enzyme. The amplified PCR fragments were then introduced into plasmid vectors PgaPZA and PVT 100L respectively and the constructs were then used to transform the for biofuel production from cocoa pods waste.
Acid hydrolysis of the pod was carried out and the engineered Saccharomyces cerevisiae was pitched into the hydrolysate at 5% (v/v) and fermentation allowed to proceed at 30oC. The fermentative characteristics of the yeast including sugar consumption, specific growth rate, Cell yield, xylose uptake rate and specific alcohol productivity were studied. With a Specific Growth rate (µ) of 0.032/hr, a specific ethanol productivity (qp) of 0.40 L/L/ hr, a final ethanol yield of 87% /L cocoa pod hydrolysate was obtained compared to 61.2%/L for the wild type. This ensures the possible use of genetically-modified yeast as enhancer of productivity in alcoholic fermentation of a cheap carbon source.
Key words: Fossilized fuel, Engineered yeast, hemicellulosic hydrolysate, Saccharomyces cerevisiae, Ethanol productivity.