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Functional Genomics and RNAi
RNAi Tuschl Protocols | ||
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Sigma has prepared product lists for commonly used protocols from Cold Spring Harbor Protocols. |
| Selected Protocols from "RNAi, A Guide to Gene Silencing", Gregory J. Hannon. Mammalian RNA Interference, Thomas Tuschl (Chapter 13, pg. 280-288) NOTE: Selected protocol links below require a subscription to Cold Spring Harbor Protocols to view. |
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| Selection of siRNA Sequences Annealing siRNAs to Produce siRNA Duplexes Cell Culture and Preparation of Cells in 24-Well Plates Cotransfection of Luciferase Reporter Plasmids with siRNA Duplexes Transfection of siRNA Duplexes Immunofluorescence Detection of Protein Knockdown Detection of Protein Knockdown by Western Blotting |
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| ABSTRACT A series of protocols is presented for targeting endogenous genes in mammalian somatic cells. |
| Products Available for "Annealing siRNAs to Produce siRNA Duplexes" |
| Protocol Material Description | Product # | Product Name | Add to Cart |
| 200 mM potassium acetate | P1190 | Potassium acetate | |
| 4 mM magnesium acetate | 63052 | Magnesium acetate solution | |
| 60 mM HEPES-KOH (pH 7.4) | H0887 | HEPES solution | |
| Ethidium bromide solution (1% w/v) aqueous | E1510 | Ethidium bromide solution | |
| Agarose suitable for small PCR fragments | A3038 | Agarose | |
| Sense and antisense siRNA in H2O at a conc. >80uM | Custom siRNA | Production of siRNA | |
| Gel loading buffer | G2526 | Gel Loading Buffer | |
| 5X TBE buffer | T6400 | Tris-Borate-EDTA buffer | |
| 450 mM Tris base | T6066 | Trizma® base | |
| 450 mM boric acid | B6768 | Boric acid | |
| 10 mM Na2EDTA | E7889 | Ethylenediaminetetraacetic acid disodium salt solution |
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| Products Available for "Cell Culture and Preparation of Cells in 24-Well Plates" |
| Protocol Material Description | Product # | Product Name | Add to Cart |
| Dulbecco's modified Eagle medium (DMEM) | D5796 | Dulbecco's Modified Eagle's Medium - high glucose | |
| Fetal bovine serum (FBS) | F2442 | Fetal Bovine Serum | |
| Penicillin and streptomycin | P4458 | Penicillin-Streptomycin | |
| Trypsin-EDTA solution | T4049 | Trypsin-EDTA solution | |
| Cell Culture flask (175 ml) | C7046 | Corning® cell culture flasks | |
| Cell culture plate (24 well) | CLS3337 | Corning® Cell Culture Plates with CellBIND® surface | |
| Coverslips | S1815 | SecureSlip™ glass coverslip |
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| Products Available for "Cotransfection of Luciferase Reporter Plasmids with siRNA Duplexes" |
| Protocol Material Description | Product # | Product Name | Add to Cart |
| Cell culture plate (24 well) | CLS3337 | Corning® Cell Culture Plates with CellBIND® surface |
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| Products Available for "Immunofluorescence Detection of Protein Knockdown" |
| Protocol Material Description | Product # | Product Name | Add to Cart |
| Hoechst 33342 | B2261 | bisBenzimide H 33342 trihydrochloride | |
| Methanol, chilled to –10°C | 494437 | Methanol | |
| 137 mM NaCl | S3014 | Sodium chloride | |
| 7 mM Na2HPO4 | S3264 | Sodium phosphate dibasic | |
| 1.5 mM KH2PO4 | 60353 | Potassium phosphate dibasic | |
| 2.7 mM KCl | P9541 | Potassium chloride | |
| Cell culture plate (24 well) carrying knockdown cells on coverslips | CLS3337 | Corning® Cell Culture Plates with CellBIND® surface | |
| Coverslips | S1815 | SecureSlip™ glass coverslip | |
| Filter paper | Z612820 | Schleicher & Schuell® filter paper | |
| Petri dish (15-cm diameter) | Z231525 | Duroplan Petri Dishes | |
| Slides | Z162701 | Corning® microscope slides, frosted one side, one end | |
| Tweezers (No. 7) | T4912 | Tweezers |
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| Products Available for "Detection of Protein Knockdown by Western Blotting" |
| Protocol Material Description | Product # | Product Name | Add to Cart |
| Blocking solution (3% milk powder in TBST [pH 7.4]) | P4739 | Phosphate buffered saline | |
| Dulbecco's modified Eagle medium (DMEM) | D5796 | Dulbecco's Modified Eagle's Medium - high glucose | |
| ECL detection kit | PQ0201 | ProteoQwest™ Chemiluminescent Western Blotting Kit, CPS Substrate | |
| Tris-Glycine Buffer | T4904 | Tris-Glycine Buffer 10× Concentrate | |
| 0.01% SDS | L4390 | Sodium dodecyl sulfate | |
| 20% methanol | 494437 | Methanol | |
| 2X Laemmli SDS sample buffer | S9788 | 2x SDS Sample Buffer | |
| 137 mM NaCl | S3014 | Sodium chloride | |
| 7 mM Na2HPO4 | S3264 | Sodium phosphate dibasic | |
| 1.5 mM KH2PO4 | 60353 | Potassium phosphate dibasic | |
| 2.7 mM KCl | P9541 | Potassium chloride | |
| Ponceau S Stain | P3504 | Ponceau S | |
| 0.2% Tween-20 | P9416 | TWEEN® 20 | |
| 20 mM Tris-HCl | T5941 | Trizma® hydrochloride | |
| 150 mM NaCl | S3014 | Sodium Chloride | |
| Trypsin-EDTA solution | T4049 | Trypsin-EDTA solution |
| back to top View all available product lists for Cold Spring Harbor Laboratory Protocols |
| Product Association Disclaimer: The Sigma-Aldrich products listed for this specific protocol were selected either to match or to supplement the products listed within the actual protocol. The products/reagents from Sigma-Aldrich have been qualified for usage, but may not have been validated for this specific application. Please refer to the detailed product description on the usage of specific products of interest. |