[PDF][PDF] Negative Regulation of MAVS-Mediated

IIR by PSMA - 2009 - researchgate.net
IIR by PSMA
2009researchgate.net
HEK293, MCF-7, and murine embryonic fibroblasts cells were grown in DMEM (Invitrogen)
supplemented with 10% heat-inactivated FBS (Hy-Clone), 2 mM L-glutamine, 100 U/ml
penicillin, and 100 mg/ml streptomycin. Transient transfections were performed with
Lipofectamine 2000 (Invitrogen) according to the manufacturer's instructions. For stable
down-regulation of PSMA7 or PSMA4, cells were transfected with pU6-PSMA7-siRNA or
pU6-PSMA4-siRNA using Lipofectamine 2000 (Invitrogen) and selected in the presence of …
HEK293, MCF-7, and murine embryonic fibroblasts cells were grown in DMEM (Invitrogen) supplemented with 10% heat-inactivated FBS (Hy-Clone), 2 mM L-glutamine, 100 U/ml penicillin, and 100 mg/ml streptomycin. Transient transfections were performed with Lipofectamine 2000 (Invitrogen) according to the manufacturer’s instructions. For stable down-regulation of PSMA7 or PSMA4, cells were transfected with pU6-PSMA7-siRNA or pU6-PSMA4-siRNA using Lipofectamine 2000 (Invitrogen) and selected in the presence of G418 3 days after transfection. G418-resistant single cell clones were amplified and screened by Western blot with anti-Flag Ab (Sigma-Aldrich). Two independent clones for each construct were harvested and used for additional experiments. The selection of the coding sequences for siRNA was based on previous guidelines. Each sequence was analyzed using the NCBI-BLAST database to verify that there was no homology greater than 15/21 residues with other genes. The sequences for the siRNA-encoding regions for PSMA7 and PSMA4 were obtained from GenBank Accession No. DQ890589 (nucleotides 324–342, http://www.
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