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Improved method for cloning DNA complementary to minor mRNAs: preparation of a hybridization probe from purified mRNAs encoding intermediate filament proteins |
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| Authors: | Philippe Brûlet Willem G. Roskam |
| Affiliation: | 1. Service de Génétique Cellulaire, Institut Pasteur, 28, rue du Docteur Roux, 75724 Paris Cedex 15 France, Tél. 33.1.306.19.19;2. Département de Génie Génétique, Institut Pasteur Production (groupe SANOF1), 3 Boulevard Raymond Poincaré, 92430 Marnes-la-Coquette France, Tél. 33.1.741.79.33, ext. 1408 |
| Abstract: | A procedure for the rapid fractionation of mRNA has been used to enrich mRNAs encoding a set of intermediate filament proteins in trophoblastoma cells. The procedure involves sucrose-gradient fractionation followed by high-resolution preparative gel electrophoresis. Part of the enriched mRNA preparation has been used to prepare a hybridization probe to screen a trophoblastoma cDNA library in Escherichia coli. A small proportion of the clones hybridized to the probe, and among these a specific clone was identified. |
| Keywords: | Recombinant DNA early embryonic differentiation preparative gel electrophoresis high resolution mRNA purification screening of cDNA clones mouse trophoblastoma cells Ac actin-specific spot cDNA DNA complementary to mRNA ds double stranded SDS sodium dodecyl sulfate TAE 40 mM Tris-acetate, 1 mM EDTA, pH 7.9 TBE 50 mM Tris-borate, 0.1 mM EDTA, pH 8.3 TCA trichloroacetic acid |
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