In order to determine the fragment size of Triticum timopheevii chromosome segments introduced into wheat background and physically map the Pm6 gene, a total of 72 primers (including SSR and STS primers) were used to analyze the eight introduced introgression lines containing Pm6 gene. Referring to the available mapping information of the analyzed markers on chromosome 2B, Pm6 was physically located in distal part of the long arm of chromosome 2B at the region of Bin 2BL-6. The introgressed fragment sizes of the chromosome 2G in different introgression lines was determined to be as follow (from short to long): IGV1-465<IGV1-464<IGV1-463<IGV1-468<(IGV1-448,IGV1-458)<IGV1-474< IGV1-466. The results were coincide with the RFLP results and further confirmed that line IGV1-465 contained the minimum introgression segment. Using the 10 polymorphism PCR markers identified in the present research, it is feasible to differentiate seven introgression lines. This information will be helpful for marker assistant identification of these lines in wheat breeding program and for further isolation of the Pm6 gene.