Abstract
The principle of mRNA purification by hybridization to an immobilized DNA fragment was applied to the isolation of mRNA coding for immunoglobulin κ-chains of mouse myeloma MOPC 21 and mouse hybridoma PTF-02. The DNA fragment comprising the 3′-untranslated region and a part of the constant region of the κ-chain gene was covalently attached to diazobenzyloxymethyl-cellulose and used as an affinity adsorbent. A homogeneous 14S mRNA species was obtained by hybridization of total mRNA to the affinity adsorbent at 52°C and by elution at 60 °C. Addition of the purified mRNA to a fractionated cell-free translation system resulted in a significant increase in the radioactivity immunoprecipitated by pig anti-mouse immunoglobulin antibodies. A single radioactive polypeptide of apparent Mr of 25,000, corresponding obviously to the κ-chain, was identified as the only translation product.
Original language | English |
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Pages (from-to) | 315-319 |
Number of pages | 5 |
Journal | Immunology Letters |
Volume | 7 |
Issue number | 6 |
DOIs | |
Publication status | Published - 1984 |
Externally published | Yes |
Keywords
- diazobenzyloxymethyl- cellulose
- immunoprecipitation
- polyacrylamide gel electrophoresis
- RNA-DNA hybridization
- translation in cell- free system
ASJC Scopus subject areas
- Immunology
- Immunology and Allergy