Nucleocápside Annealing-Mediated Eletroforese (NAME) ensaio permite a identificação rápida de HIV-1 Nucleocápside Inibidores
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Summary
Abstract
Introduction
Protocol
Representative Results
Discussion
Disclosures
The authors have nothing to disclose.
Acknowledgements
Materials
| cTAR, 29-mer DNA oligo. Desalted. HPLC purified. | Metabion International AG | Store the stock solution at -20 °C | |
| TAR, 29-mer RNA oligo. Desalted. HPLC purified. | Metabion International AG | Store the stock aliquot at -80 °C | |
| (12-55)NC peptide | EspiKem srl | Store the stock solution at -20 °C | |
| 1.5 ml tubes | Eppendorf | 0030 120 086 | Autoclave before use |
| 0.5 ml tubes | Eppendorf | 0030 121 023 | Autoclave before use |
| Biosphere Filter Tips 0.1-10 µl | Sarstedt | 701,130,210 | |
| Biosphere Filter Tips 2-20 µl | Sarstedt | 70,760,213 | |
| Biosphere Filter Tips 200 µl | Sarstedt | 70,760,213 | |
| Syringe Filter 0.22 µm | Biosigma srl | 51,798 | |
| Ethanol | Carlo Erba | 414631 | |
| Sodium Dodecyl Sulfate | Sigma-Aldrich | 71725-50G | |
| Dimethyl Sulfoxide | Sigma-Aldrich | D8418-1L | |
| Sodium Chloride | Sigma-Aldrich | 71376-1KG | |
| Boric Acid | Sigma-Aldrich | B0252-2.5KG | |
| Ethylenediaminetetraacetic Acid | Sigma-Aldrich | E5134-500G | |
| Magnesium Perchlorate | Sigma-Aldrich | 222283-100G | Store the 1 M solution at -20 °C |
| Glycerol | Sigma-Aldrich | 49770-1L | |
| Bromophenol Blue | GE Healthcare Life Sciences | 17-1329-01 | |
| <em>N,N,N′,N′</em>-Tetramethylethylenediamine | Sigma-Aldrich | T9281-100ML | |
| Ammonium Persulfate | Sigma-Aldrich | A7460-500G | Prepare the 10% solution freshly before use |
| Acrylamide-bis ready-to-use solution 40% (19:1) | Merck Millipore | 1006401000 | Polyacrylamide is highly neurotoxic: wear gloves during the gel casting |
| Tris ultrapure | Applichem | A1086,1000 | |
| DEPC-treated water | Ambion | AM9922 | |
| Centrifuge 5415R | Eppendorf | 22,621,408 | |
| NanoDrop 1000 spectrometer | Thermo Scientific | ||
| UV-VIS spectrometer Lambda 20 | Perkin Elmer | ||
| Protean II xi Cell | Bio-Rad | 165-1803 | |
| PowerPac Basic Power Supply | Bio-Rad | 164-5050 | |
| SybrGreen II RNA Gel Staining | Lonza | 50523 | Make the 1/10,000 dilution in TBE 1x. Store it in the dark at 4 °C for two weeks. |
| Geliance 600 Imaging System | Perkin Elmer |
References
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Cite This Article
Sosic, A., Cappellini, M., Scalabrin, M., Gatto, B. Nucleocapsid Annealing-Mediated Electrophoresis (NAME) Assay Allows the Rapid Identification of HIV-1 Nucleocapsid Inhibitors. J. Vis. Exp. (95), e52474, doi:10.3791/52474 (2015).