Our main concerns were whether i) the thermotolerant mutants exhibited improved ethanol yield, and ii) the mutants were applied to stimultaneous saccharification and fermentation with pretreated Miscanthus at high temperature. Thus, the four strains were further screened by their ability to grow and to ferment glucose in synthetic broth at 42 C. As expected, the overall growth rate of the mutants and ethanol concentration were distinctly higher than those of the control at 42 C (Fig.3). The mutants produced 9.7 g/L ethanol on average at 42 C, which was 2.2-fold higher than the ethanol concentration produced by the control. The maximal ethanol concentration was 10.2 g/L, achieved by the S.cerevisiae mbc 2 strain (Fig.3-v). Thus, S.cerevisiae mbc 2 among the mutants finally was selected as a suitable strain for bioethanol production at high temperature. However, the average ethanol yield of the mutants was far lower than the theoretical yield, possibly because of the lower initial cell density, which was adjusted to OD600 = 0.2 for high temperature. To overcome this limitation, high-cell-density fermentation should be further considered for SSF [38]. The S.cerevisiae mbc 1 strain was shown to have different growth modalities on broth and solid cultures for unexplained reasons.However, this difference may be attributed to environmental differences.(e.g,. differences in diffusion in solid and broth medium). Kim et al. [22] also encountered this difference which was not fully explained. Meanwhile, at 30 C, the growth rate of all thermotolerant mutants was much lower than that of the control over a 12-h period when the ethanol concentration of all strains reached an average maximum of 21.6 g/L (Fig. 3-a). This phenomenon may be explained by a trade-off between stress tolerant acquisition and growth rate under a non-stressful environment.[39]