TranslationBlocker Human and Mouse Apoptosis regulator Bcl-2 siRNA – DISCONTINUED

DISCONTINUED: Human and Mouse Apoptosis regulator Bcl-2 TranslationBlocker™ siRNA Duplex Product Attributes

Apoptosis regulator Bcl-2 Knockdown Protocol:

1. 1. Starting with cultured cells in growth media seed 6 well plates with 1.5×10⁵ cells and incubate at 37°C for 24 hours.
   2. Prepare transfection reagent in serum-free growth media such as Opti-MEM allow to come to room temperature. The above data was generated with Oligofectamine for delivery of human / mouse Bcl-2 siRNA prepared in accordance with manufacturer’s instructions.
   3. Dilute siRNA to working concentration in serum-free growth media. For Oligofectamine or similar transfections into 6 well plates, we recommend starting siRNA concentration optimization at around 100 pmol siRNA duplex in 750 ul of serum free media. Optimal dilution should be experimentally determined.
   4. Combine diluted siRNA and transfection reagent. Equivalent volumes of prepared Oligofectamine or other transfection reagent and diluted siRNA should be combined and incubated at room temperature for 30 minutes with gentle mixing (optional).
   5. Add prepared siRNA solution to cells. Additionally, FITC labeled control siRNA (Product Number QC-1F) or positive control siRNA (Beta-Actin; Product Number QC-2) may be used to assess transfection efficiency.
   6. Incubate for several hours in serum free media.
   7. Add serum containing media to culture without a media change. Generally 1-2 fold of the original volume of siRNA containing serum free media is added to restore nutrient levels in media.
   8. Incubate and assay several hours after transfection. Optimal knockdown time should also be experimentally determined. Optionally, a second transfection can be performed 24 hours after the first transfection to enhance knockdown. Human Bcl-2 knockdown at the protein level was measured at 24, 48, and 72 hours post transfection via western blot.