Abstract: Mu Opioid receptor plays an important role in mediating most of diverse effects of opioids like analgesia, euphoria and dependence. Studies on Glycosylation Consensus of various receptors showed that it has important roles in the affinity, desensitization, G-protein coupling and intracellular signaling. The aim of the present study was to create a mutation in this consensus of Rat Mu Opioid Receptor (RMOR) for further investigation of its function. Using nested PCR one mutation was produced, N53Q. The PCR products as well as the pcDNA3 vector were digested using HindIII and BamHI restriction enzymes and were ligated and transformed to Ecoli HB101 cells. The obtained colonies were analyzed for the presence of the inserts using suitable restriction enzymes. The obtained plasmids are ready for further investigations in binding assay and function assays like cyclic AMP measurements.