Abstract: Uredospores of wheat yellow rust was used as test material. Isolation methods of wheat yellow rust genomic DNA by CTAB，SDS，Urea and CTAB/SDS were compared．The results showed that the purity and yield of wheat yellow rust DNA obtained by above methods were different．These four methods were ranked as: CTAB＞Urea＞CTAB/SDS＞SDS according to the average DNA microgram of uredospores, and ranked as：CTAB/SDS＞CTAB＞SDS＞Urea according to the genomic DNA purity．The DNA obtained by these four methods was subjected to SSR analyses with RJ17 primer ,and the DNA obtained with CTAB method used as template was better than others in SSR analysis．