HTRA2 MISSION shRNA Lentiviral Transduction Particles HtrA serine peptidase 2

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	To establish a stable HtrA2/Omi-depleted cell line, FL83B cells were infected with a mouse HtrA2/Omi specific shRNA-encoded lentivirus (Sigma; SHCLNV-NM_019752). An shRNA negative control lentiviral particle (LV-Control) was used as a negative control. To generate a stable cell line, FL83B cells were plated at a density of 1 × 105 cells per 60-mm culture dish and infected overnight with five multiplicities of infection (MOI) lentiviral particles in the presence of 8 μg/mL hexadimethrine bromide (Sigma). After infection, the transduced cells were selected using 10 mg/mL puromycin (Sigma) for 2 weeks and incubated at 37 °C in a humidified incubator with 5% CO2. Suppression of HtrA2/Omi expression in selected cells was confirmed by western blot analysis.

Protocol tips

To establish a stable HtrA2/Omi-depleted cell line, FL83B cells were infected with a mouse HtrA2/Omi specific shRNA-encoded lentivirus (Sigma; SHCLNV-NM_019752). An shRNA negative control lentiviral particle (LV-Control) was used as a negative control. To generate a stable cell line, FL83B cells were plated at a density of 1 × 105 cells per 60-mm culture dish and infected overnight with five multiplicities of infection (MOI) lentiviral particles in the presence of 8 μg/mL hexadimethrine bromide (Sigma). After infection, the transduced cells were selected using 10 mg/mL puromycin (Sigma) for 2 weeks and incubated at 37 °C in a humidified incubator with 5% CO2. Suppression of HtrA2/Omi expression in selected cells was confirmed by western blot analysis.

Publication protocol

To establish a stable HtrA2/Omi-depleted cell line, FL83B cells were infected with a mouse HtrA2/Omi specific shRNA-encoded lentivirus (Sigma; SHCLNV-NM_019752). An shRNA negative control lentiviral particle (LV-Control) was used as a negative control. To generate a stable cell line, FL83B cells were plated at a density of 1 × 105 cells per 60-mm culture dish and infected overnight with five multiplicities of infection (MOI) lentiviral particles in the presence of 8 μg/mL hexadimethrine bromide (Sigma). After infection, the transduced cells were selected using 10 mg/mL puromycin (Sigma) for 2 weeks and incubated at 37 °C in a humidified incubator with 5% CO2. Suppression of HtrA2/Omi expression in selected cells was confirmed by western blot analysis.

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