Objective：To investigate microRNA-29b (miR-29b) expression in cerebral cortex, spinal cord, fore limb muscle, and serum of SOD1-G93A amyotrophic lateral sclerosis (ALS) mice, and to identify the biomarker and to assess diagnostic values for ALS. Methods: Cerebral cortex, spinal cord, fore limb muscle and serum from 16 SOD1-G93A ALS mice and 16 wild-type mice were taken and then microRNA extracted, detecting the expression of miR-29b by real-time quantitative polymerase chain reaction (RT-qPCR). The diagnostic performance of miR-29b for ALS was estimated by the receiver operating characteristic (ROC) curve. Results: The results from the validation indicated that the differences in miR-29b between the cerebral cortex of SOD1-G93A ALS and the healthy control subjects were statistically significant (P=0.001). Meanwhile, the expressions 8, 12, and 16 weeks later were higher than those of the controls (ALS vs. Control: 8 weeks, P=0.044; 12 weeks, P=0.018; 16 weeks, P=0.045). When the relative expression level of miR-29b was used to diagnose ALS in SOD1-G93A ALS mice, the area under the ROC (area under the curve, AUC) was 0.885, if the diagnostic threshold was set at 0.185 6, the sensitivity and specificity were 92.9% and 71.4%. Conclusion: MiR-29b may act as medical monitoring indices of ALS in early time.