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ABSTRACT
Chromatography on oligo(dT) columns is the preferred method for large-scale purification (>25 mg) of poly(A)+ RNAextracted from mammalian cells. Typically, between 1% and 10% of the RNA applied to the oligo(dT) column is recovered as poly(A)+ RNA. Because the method can be frustratingly slow, it is not recommended for purification of poly(A)+ RNA from multiple samples. For this purpose, batch elution (Protocol 7.4) is the better choice.
IMPORTANT: Prepare all reagents used in this protocol with DEPC-treated H2O.
Products Available for this Protocol
| Protocol Material Description |
Product # |
Product Name |
Add to Cart |
| 2x Column-loading buffer |
|
|
|
| 40 mM Tris-HCl (pH 7.6) |
T5941 |
Trizma® hydrochloride for molecular biology, Biotechnology Performance Certified |
|
| 1 mM NaCl |
S3014 |
Sodium chloride for molecular biology |
|
| 2 mM EDTA (pH 8.0) |
E5134 |
Ethylenediaminetetraacetic acid disodium salt solution for molecular biology |
|
| Elution Buffer |
|
|
|
| 10 mM Tris-HCl (pH 7.6) |
T5941 |
Trizma® hydrochloride for molecular biology, Biotechnology Performance Certified |
|
| 1 mM EDTA (pH 8.0) |
E5134 |
Ethylenediaminetetraacetic acid disodium salt solution for molecular biology |
|
| 0.05% SDS |
L4390 |
Sodium dodecyl sulfate for molecular biology |
|
| NaCl (5M), Rnase-free |
S3014 |
Sodium chloride for molecular biology |
|
| Ethanol |
E7023 |
Ethanol |
|
| NaOH (10 N) |
72068 |
Sodium hydroxide solution, BioChemika Ultra, for molecular biology |
|
| Sodium acetate (3 M, pH 5.2) |
S7899 |
Sodium acetate buffer solution |
|
| Oligo(dt)-cellulose |
O3131 |
Oligo(dT)-Cellulose |
|
| pH Paper (pH Test Strips) |
P4786 |
pH Test strips |
|
Note: As an alternative, the entire protocol could be accomplished with product MRN10 or MRN70, the GenElute™ mRNA Miniprep Kits.
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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.
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