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Stem rust, caused by Puccinia graminis f. sp. tritici (Pgt), is one of the most important diseases of wheat with devastating epidemics in Iran and the world. In this study, we evaluated some Iranian wheat landraces in a greenhouse at the seedling stage against a new pathotype related to Ug99 of Pgt, which was collected from Iran and designated as TTSSK. Marker analysis was done on resistant landraces. Molecular markers for detecting some Sr genes were used. The results showed that Sr22, Sr35 and SrWeb provided resistance against TTSSK in the resistant landraces. In addition, some of the susceptible landraces that were resistant at adult stage were used for Sr2 analysis. The results showed that some of these landraces were carrying other adult plant resistance gene/genes except Sr2. To evaluate the defence gene expression in compatible and incompatible interactions, cv. Morocco (susceptible) and KC-440 landrace (resistant) were used. Sampling was done at 0, 12, 18, 24, and 72 hours post inoculation (hpi) with stem rust isolate and water as mock treatment. β-1,3 glucanase gene expressions were studied using qGLU-S and qGLU-AS primers. Also, 18srRNA, β-tubulin and EF1-α genes were used as internal control. The results showed that in incompatible interactions, the defence gene expression was increased at 24 hpi, but in compatible interactions, expression level reached the peak at 12 hpi and it significantly decreased at 18 hpi. The results revealed that the expression of defence genes such as β-1,3 glucanase was earlier in compatible interactions than in incompatible interactions, but the quantity of expressed gene was less than in incompatible interactions. [ABSTRACT FROM AUTHOR]