Anti mor antibody

Stable Control and MOR shRNA (Santa Cruz Biotechnology, Santa Cruz, CA) were stably transfected into H358 cells as we have previously described [12] (link). Cells (∼40% confluent) were serum-starved for 1 hour followed by incubated with shRNA for 6 hours in serum-free media. Serum-containing media was then added (10% serum final concentration) for 42 hours and puromycin selection reagent was added. Inhibition of protein expression was confirmed by immunoblot analysis with anti-MOR antibody (GeneTex, San Antonio, TX).

Myc-DDK-tagged ORF clone of Homo sapiens opioid receptor, mu 1 (OPRM1), transcript variant MOR-1 (OriGene Technologies Inc, MD) was amplified using Platinum Taq DNA polymerase high fidelity enzyme (Invitrogen, CA) and subsequently cloned into a pCR8/GW/Topo entry vector (Invitrogen, CA) according to manufacturer's instructions. Plasmid DNA was extracted from selected clones by QIAquick Plasmid Mini kit (Qiagen, CA). ORF integrity and fragment orientation were confirmed by sequencing. The MOR1-Myc fusion product was then transferred to pcDNA3.2/v5 DEST vector (Invitrogen, CA) by LR reaction. The resulting construct (pcDNA3.2-MOR1-Myc) was transfected into H358 cells using FuGENE HD™ as the transfection reagent (Roche Applied Sciences) according to the protocol provided by Roche as we have previously described. Cells (∼40% confluent) were serum-starved for 1 hour followed by incubation with pcDNA3.2-MOR1-Myc for 6 hours in serum-free media. Serum-containing media was then added (10% serum final concentration) for 42 hours and neomycin selection reagent was added. Overexpression was confirmed by immunoblot analysis with anti-MOR antibody (GeneTex, San Antonio, TX).