DISCONTINUED: Human TLR4 TranslationBlockerâ¢ siRNA Duplex Product Attributes
Blocks transcription and translation of TLR4 from: Human.
May also react with TLR4 based upon sequence homology and clone specific publications.
Type of RNAi: siRNA.
siRNA target sequence: NM_138554.4.
Samples and cell lines tested: HepG2 Human Carcinoma Cells from Liver.
Buffer and Stabilizer: Lyophilized. See datasheet for resuspension instructions and buffer information.
Concentration of Anti-Human TLR4: Lyophilized
SiRNA Purification Method:RPC/HPLC
Storage Conditions: Store Lyophilized siRNA duplex at 4C. Reconsituted product may be stored at -20C for up to 1 month.TLR4 Knockdown Protocol:
- Starting with cultured cells in growth media seed plates with cells and incubated at 37Â°C. The above data was generated in 6 well plates with 5x105 cells incubated at 37Â°C until cells reached 70% confluence prior to starting transfections
- Prepare transfection reagent in serum-free growth medium such as Opti-MEM-I. Allow to come to room temperature. Lipofectamine 2000 was used to generate the above data in accordance with manufacturerÂs instructions.</ li>
- Dilute siRNA to working concentration in serum-free growth media. For Lipofectamine or similar transfections into 6 well plates, we recommend starting siRNA dose optimization at around 150 pmol siRNA duplex.Optimal dilution should be experimentally determined.
- Combine diluted siRNA and transfection reagent. Ratio of transfection reagent to siRNA should be optimized. Equivalent volumes of prepared Lipofectamine 2000 or other transfection reagent and diluted siRNA should be combined and incubated at room temperature for 30 minutes with gentle mixing (optional).
- Add prepared siRNA solution to cells. For 6 well plates this will entail adding the solution to cells in approximately 2.5 ml serum-free medium. Optionally, 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.
- Incubate for several hours in serum free media. The above data was generated with a 24 hour incubation.
- Add serum containing media to culture without a media change if longer incubation times are required. Generally 1-2 fold of the original volume of siRNA containing serum-free media is added to restore nutrient levels in media. Alternatively, the media may be directly supplemented with serum such as 10% FBS.
- 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. TLR4 knockdown at the protein level was measured at 48 hours post transfection via Western Blot.
TLR4 General Information
Gene Name: TLR4
Entrez Gene ID: 7099
NCBI Ref Seq Accession Number: NP_612564
TLR4 Molecular Weight:
Alternate Names: TLR-4, Toll-like Receptor 4, CD284
- Pan, J., Huang, H., Sun, L., Fang, B. & Yeung, S.-C. J. Bcl-2-associated X protein is the main mediator of manumycin a-induced apoptosis in anaplastic thyroid cancer cells. J. Clin. Endocrinol. Metab. 90, 3583Â91 (2005).
- BidÃ¨re, N. et al. Cathepsin D triggers Bax activation, resulting in selective apoptosis-inducing factor (AIF) relocation in T lymphocytes entering the early commitment phase to apoptosis. J. Biol. Chem. 278, 31401Â11 (2003).
Limitations and Warranty
This product is for Research Use Only. This product is guaranteed to work for a period of two years when stored at -70C or colder, and one year when aliquoted and stored at -20C.