Abstract
During the last few years, the global transcription machinery engineering (gTME) technique has gained more attention as an effective approach for the construction of novel mutants. Genetic strategies (molecular biology methods) were utilized to get mutational for both genes (SPT15 and TAF23) basically existed in the Saccharomyces cerevisiae genome via screening the gTME approach in order to obtain a new mutant S. cerevisiae diploid strain. The vector pYX212 was utilized to transform these genes into the control diploid strain S. cerevisiae through the process of mating between haploids control strains S. cerevisiae (MAT-a [CICC 1374]) and (MAT-α [CICC 31144]), by using the oligonucleotide primers SPT15-EcoRI-FW/SPT15-SalI-RV and TAF23-SalI-FW/TAF23-NheI-RV, respectively. The resultant mutants were examined for a series of stability tests. This study showed how strong the effect of using strategic gTME with the importance of the modification we conducted in Error Prone PCR protocol by increasing MnCl2 concentration instead of MgCl2. More than ninety mutants we obtained in this study were distinguished by a high level production of bio-ethanol as compared to the diploid control strain.
Highlights
Ethanol (ETOH) is a colorless liquid produced by the fermentation of sugars by a variety of microbes such as Saccharomyces cerevisiae (S. cerevisiae), Escherichia coli (E. coli), Klebsiella oxytoca and Zymomonas mobilis (Tan et al 2016)
Up to the present time, the global transcription machinery engineering (gTME) technique has been used for the construction of new mutant; in our study we processed a new yeast strain for a library of novel mutants by using SPT15 gene
The gTME technique has been modified by using M nCl2 instead of the most applied mixture Ep-polymerase chain reaction (PCR) reaction MgCl2
Summary
Ethanol (ETOH) is a colorless liquid produced by the fermentation of sugars by a variety of microbes such as Saccharomyces cerevisiae (S. cerevisiae), Escherichia coli (E. coli), Klebsiella oxytoca and Zymomonas mobilis (Tan et al 2016). It is used as fuel, solvent, sterilizer, antiseptic for wounds, cosmetics and pharmaceutical industry. Ep-PCR is a fast and cheap molecular biology method for random mutation in a particular piece of DNA. This technique is a based on PCR and its reaction mixture composition.
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