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Weeds such as wild mustard are a serious problem in crop production. Herbicide resistance in weeds and wild mustard has become a common threat in agriculture and endangers global food security. In this experiment, the ALS gene of wild weed suspected of weeding due to field screening and PCR-based methods from Kermanshah province's wheat fields was investigated. The ALS gene was then identified, isolated and cloned. For this purpose, primers were designed according to the information available in the NCBI database and ALS gene was amplified using RT-PCR. The gene ALS cloned in pTZ57R / T vector within E. coli bacteria, DH5α strain and sequenced. Based on PCR-based molecular results, six biotypes showed mutations in the target site. PCR-based sequencing results confirm the nucleotide mutations of Asp376-Gly (GAC to GGC), Pro197-Ser (CCT to TCT), Trp574-Leu (TGG to TTG), and Ala122-Thr (GCA to ACA) in six biotypes. These mutation sites were confirmed by sequencing. According to the study, herbicide resistance in wild mustard could be related to resistance in the target site due to the mentioned nucleotide mutations in the active site of the enzyme, the binding site of ALS inhibitors and its family, or due to the resistance of the target site that can cause conformational changes. Changes in the level of the protein reaction site with herbicides and as a result changes in activity and enzyme-herbicide binding. This means that the molecular levels of the mutant enzyme are less likely to come into contact with herbicides, and the herbicide cannot stop the enzyme from working and cause herbicide resistance.

Keywords: Nucleotide mutation, PCR based method, herbicide resistance, ALS inhibitor, wild mustard

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