Genetical engineering techniques have been applied to increase substrate range in microorganisms such as S. cerevisiae and Z. mobilis that help to maximize ethanol production like that in E. Coli. It also supplies other important traits for conversion of lignocellulose into ethanol. Since the molecular basis for ethanol and inhibitor tolerance is not fully understood, random mutagenesis and evolutionary engi­neering have also been applied to improve those traits. Moreover, as a result of technological developments, systems biology approaches have recently been applied to characterize the functional genomics of microorganisms and to evaluate the impact of metabolic and evolutionary engineering strategies. This advanced characterization (genomics, transcriptomics, proteomics, metabolomics) is already contributing to better understand that physiological responses and to identify crucial targets for metabolic engineering [14, 90, 189].