Gut microbiota plays an important role in the occurrence and development of type 2 diabetes mellitus. In order to further explore the effect of foxtail millet on gut microbiota of diabetic rats, the 16S rRNA high throughput sequencing technology was used to compare the changes of gut microbial composition and relative abundance in the rats fed with high-fat diet combined with STZ, from two dimensions of horizontal (different groups) and vertical (before and after intervention), respectively. It was shown that foxtail millet can significantly increase the relative abundance of Ruminococcus_2 and Lactobacillus, and significantly reduce the relative abundance of Blautia. Foxtail millet intervention also decreased the ratio of Bacteroides to Firmicum in high-fat-diet and STZ induced diabetic rats. And these changes were related to physiological indexes. In conclusion, foxtail millet intervention can reverse the effect of type 2 diabetes on gut microbiota, and make the gut microbial composition of rats develop toward a more healthier direction.