Abstract: Three goats which fisted with permanent rumen cannulas were used to provide rumen fluid to harvest microbes in vitro, and to investigate the effects of rations in different starch to cellulose ratio on recovery rate of microbes from rumen fluid after detaching and rate of DNA extraction from rumen microbes. Substrates were designed by varying the ratio of starch to cellulose (NSC∶SC) as follows: 100∶0, 70∶30, 50∶50, 30∶70, 0∶100. Results showed that the recovery rate of microbes from rumen fluid after detaching and the rate of DNA extraction from rumen microbes generally decreased with the increase of dietary cellulose ratio. The average recovery rate of microbes was 53.29%, and the average rate of DNA extraction from mixed microbes was 51.0%, it was further observed that the rate of DNA extraction from bacteria (45.4%) were significantly less than those samples from protozoa (56.1%) (P<0.05). The results also showed that the molecular size of DNA fragment were larger than 20 kb, and the DNA which extracted from all the samples were fit for further studies.