Total RNA Isolation Kit TRIzol , Total RNA Purification Kit From Cultured Animal Cells

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Total RNA Purification Kit(TRIzol) Offered in twosizes (25preps &50 preps), RP2401&RP2402 Biotekes Total RNA Purification Kit provides a rapid method for the isolation and purification of total RNA from cultured animal cells, tissue samples, blood, plasma, serum, bacteria, yeast, fungi, plants and viruses. The kit purifies all sizes of RNA, from large mRNA and ribosomal RNA down to microRNA (miRNA) and small interfering RNA (siRNA). The RNA is preferentially purified from other cellular components such as proteins, without the use of phenol or chloroform. The purified RNA is of the highest integrity, and can be used in a number of downstream applications including real time PCR, reverse transcription PCR, Northern blotting, RNase protection and primer extension, and expression array assays. Biotekes Purification Technology Purification is based on spin column chromatography using Biotekes proprietary resin as the separation matrix. The RNA is preferentially purified from other cellular components such as proteins without the use of phenol or chloroform. The process involves first lysing the cells or tissue of interest with the provided Buffer RL (please see the flow chart on page 4). Ethanol is then added to the lysate, and the solution is loaded onto a spin-column. Biotekes resin binds RNA in a manner that depends on ionic concentrations. Thus only the RNA will bind to the column, while the contaminating proteins will be removed in the flowthrough or retained on the top of the resin. The bound RNA is then washed with the provided Wash Solution A in order to remove any remaining impurities, and the purified total RNA is eluted with the Elution Solution A. The purified RNA is of the highest integrity, and can be used in a number of downstream applications. Storage Conditions and Product Stability All solutions should be kept tightly sealed and stored at room temperature. These reagents should remain stable for at least 1 year in their unopened containers. Advantages # Fast and easy processing using rapid spin-column format # Isolate total RNA, from large rRNA down to microRNA (miRNA) # No phenol or chloroform extractions # Isolate high quality total RNA from a variety of sources # RNA can be isolated and detected from as little as a single animal cell Precautions and Disclaimers This kit is designed for research purposes only. It is not intended for human or diagnostic use. Ensure that a suitable lab coat, disposable gloves and protective goggles are worn when working with chemicals. The Buffer RL contains guanidinium salts, and should be handled with care. Guanidinium salts form highly reactive compounds when combined with bleach, thus care must be taken to properly dispose of any of these solutions Blood of all human and animal subjects is considered potentially infectious. All necessary precautions recommended by the appropriate authorities in the country of use should be taken when working with whole blood. Customer-Supplied Reagents and Equipment You must have the following in order to use the Total RNA Purification Kit: For All Protocols # Benchtop microcentrifuge # 96 - 100% ethanol # -mercaptoethanol (optional) For Animal Cell Protocol PBS (RNase-free) For Animal Tissue Protocol # Liquid nitrogen # Mortar and pestle # 70% ethanol For Nasal or Throat Swabs Sterile, single-use cotton swabs For Bacterial Protocol Lysozyme-containing TE Buffer: o For Gram-negative bacteria, 1 mg/mL lysozyme in TE Buffer o For Gram-positive bacteria, 3 mg/mL lysozyme in TE Buffer For Yeast Protocol Resuspension Buffer with Lyticase: o 50 mM Tris pH 7.5 o 10 mM EDTA o 1 M Sorbital o 1 unit/L Lyticase For Fungi Protocol # Liquid nitrogen # Mortar and pestle # 70% ethanol For Plant Protocol # Liquid nitrogen # Mortar and pestle # 70% ethanol For Plasma/Serum Protocol MS2 RNA (0.8 g/l). (Roche, Cat. No. 10165948001) Working with RNA RNases are very stable and robust enzymes that degrade RNA. Autoclaving solutions and glassware is not always sufficient to actively remove these enzymes. The first step when preparing to work with RNA is to create an RNase-free environment. The following precautions are recommended as your best defense against these enzymes. # The RNA area should be located away from microbiological work stations # Clean, disposable gloves should be worn at all times when handling reagents, samples, pipettes, disposable tubes, etc. It is recommended that gloves are changed frequently to avoid contamination # There should be designated solutions, tips, tubes, lab coats, pipettes, etc. for RNA only # All RNA solutions should be prepared using at least 0.05% DEPC-treated autoclaved water or molecular biology grade nuclease-free water # Clean all surfaces with commercially available RNase decontamination solutions # When working with purified RNA samples, ensure that they remain on ice during downstream applications

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