Bioethanol is considered the most proposed next automotive fuel generation. For the bioethanol production, the microbial conversion from some local environmental wastes (rice straw and manure) and the indigenous strains, namely Streptomyces aegyptia and Candida tropicalis were used in this study. Improving the conditions that may affect the process such as alkali pretreatment, enzymatic hydrolysis, and fermentation were investigated using the Plackett-Burman design (PBD) via submerged fermentation. X-ray diffraction (XRD) analysis revealed that the crystallinity index of pretreated rice straw (52.2%) was significantly (p-value) lower as compared to the untreated one (63.1%). The results showed that the optimization of the microbial conversion reached 6.6 folds in comparison to the unoptimized process. An improvement in enzymatic activities of Streptomyces aegyptia was achieved in the design that the maximal lignin peroxidase, Mn-dependent peroxidase, total cellulase, endocellulase, and xylanase activities were 120 U/min.l, 1020 U/min.l, 3.7 U/min.l, 57.3 U/min.l and 78.1 U/min.l, respectively. Maximum ethanol concentration and bioconversion process efficiency were 0.8 g/l and 13.3%, respectively.