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Invitrogen™ Acid-Phenol:Chloroform, pH 4.5 (with IAA, 125:24:1)
Description
Acid Phenol:Chloroform:IAA (125:24:1) is premixed and supplied at pH 4.5 ± 0.2. Provided in one bottle of 400 mL. In RNA extraction procedures, Acid Phenol:Chloroform:IAA aids in the removal of DNA (it partitions into the organic phase), helps to stabilize the interface, and prevents foaming when mixing. Preparation of phenol for use in molecular biology applications is a time-consuming and often hazardous procedure due to its toxic and corrosive nature. Ambion premixed, quality-tested, saturated phenols are ready to use, eliminating handling problems and offering a more convenient, safer, and easier alternative to preparing solutions from crystalline phenol.
Order Info
Shipping Condition: Room temperature
Specifications
Specifications
| Chemical Name or Material | Phenols |
| Content And Storage | Store at 4°C or -20°C. |
| Packaging Type | Bottle |
| pH | 4.5 |
| Product Line | Ambion |
| Purity | Molecular Biology Grade |
| Quantity | 100 mL |
| Shipping Condition | Room Temperature |
| Form | Liquid |
Frequently Asked Questions (FAQs)
Phenol extraction of proteins:
Phenol extraction is frequently used to remove proteins from nucleic acid solutions. A common protocol is to add an equal volume of buffer-saturated phenol or phenol:chloroform:isoamyl alcohol (25:24:1, v/v/v) to an aqueous nucleic acid solution, vortex, and centrifuge at 14,000 x g for 1 min to separate the phases.
Studies at Thermo Fisher Scientific have shown that the concentration of NaCl in the aqueous solution should not exceed 0.5 M for good recovery of DNA. Residual phenol can be removed from the aqueous phase by extraction with an equal volume of chloroform or ether. After extraction, DNA is usually precipitated with ammonium acetate and ethanol as described in another protocol on this server. Ref. Karger, B. D. (1989) FOCUS 11, 14.
A good source of general information on the properties of phenol can be found in Wallace, Donald M. “Large and Small-Scale Phenol Extractions”. Methods in Enz. Volume 152 guide to Molecular Cloning Techniques. 1987. Academic Press, Inc. Berger and Kimmel, eds. Chap.4, pg 33-41.
(a) At pH 5 to 6 DNA is selectively retained in the organic phase and interphase, leaving RNA in the aqueous phase. Therefore a pH greater than 7 is needed if DNA is to be extracted.
(b) At pH values below 7.6, poly A+ RNA is lost to the organic phase if chloroform is not present.
(c) Optimal RNA yields in phenol extraction are obtained if the salt concentration is less than 0.15 M NaCl. Salt concentration in the sample is not a factor for larger DNA molecules.
To store RNA after extraction use DEPC-treated water.
Below is a commonly used protocol:
(1) Add an equal volume of buffer-saturated phenol or phenol:chloroform:isoamyl alcohol (25:24:1, v/v/v) to an aqueous nucleic acid solution. Note: for RNA solutions, acid-phenol is recommended.
(2) Vortex, and centrifuge at 14,000 x g for 1 min to separate the phases.
(3) Residual phenol can be removed from the aqueous phase by extraction with an equal volume of chloroform or ether.
(4) After extraction, DNA/RNA is usually precipitated with ammonium acetate and ethanol.
Partitioning of the nucleic acids in phenol is pH dependent. At pH 7.0 or higher, both DNA and RNA partition into the aqueous phase. At an acidic pH (below 7.0) DNA is denatured and will move into the organic phase, but the RNA remains in the aqueous phase. The mixture should be adjusted to at least pH 7.4 for work with DNA.
Safety and Handling
For Research Use Only. Not for use in diagnostic procedures.
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