A novel constructed SPT15 mutagenesis library of Saccharomyces cerevisiae by using gTME technique for enhanced ethanol production
Ashraf A. M. M. El-Rotail,
Liang Zhang,
Youran Li,
Shuang Ping Liu,
Gui Yang Shi
Affiliations
Ashraf A. M. M. El-Rotail
The Key Laboratory of Industrial Biotechnology, Ministry of Education, National Engineering Laboratory for Cereal Fermentation Technology, School of Biotechnology, Jiangnan University
Liang Zhang
The Key Laboratory of Industrial Biotechnology, Ministry of Education, National Engineering Laboratory for Cereal Fermentation Technology, School of Biotechnology, Jiangnan University
Youran Li
The Key Laboratory of Industrial Biotechnology, Ministry of Education, National Engineering Laboratory for Cereal Fermentation Technology, School of Biotechnology, Jiangnan University
Shuang Ping Liu
The Key Laboratory of Industrial Biotechnology, Ministry of Education, National Engineering Laboratory for Cereal Fermentation Technology, School of Biotechnology, Jiangnan University
Gui Yang Shi
The Key Laboratory of Industrial Biotechnology, Ministry of Education, National Engineering Laboratory for Cereal Fermentation Technology, School of Biotechnology, Jiangnan University
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.
