Isolation and characterization of IFN-gamma mRNA derived from mitogen-induced human splenocytes.
Human splenocytes from large cultures (5 1) were stimulated with Staphylococcus enterotoxin A (SEA). After 2.5 d of conditioning, poly A+RNA was isolated from these cultures and its in-vitro translational capacity was evaluated in a wheat germ extract. Injection of the RNA into Xenopus laevis oocytes yielded a low but reproducible amount of IFN activity, which was neutralizable only by an antiserum to gamma interferon (IFN-gamma) and not by antisera to alpha interferon (IFN-alpha) and/or beta interferon (IFN-beta). Formamide sucrose gradient centrifugation analysis indicated that the mRNA which codes for IFN-gamma sediments at around 15 S. The profile, however, suggested a size heterogeneity of IFN-specific mRNA. Electrophoresis of cDNA synthesized on these mRNA fractions indicated that the mRNA coding for gamma IFN has a length of between 900 and 1400 nucleotides.
D000818:Animals; D002478:Cells, Cultured; D004768:Enterotoxins / Q000494:pharmacology; D005260:Female; D006801:Humans; D066298:In Vitro Techniques; D007371:Interferon-gamma / Q000235:genetics*; D008934:Mitogens / Q000494:pharmacology; D009865:Oocytes / Q000378:metabolism; D011061:Poly A / Q000378:metabolism; D014176:Protein Biosynthesis; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000302:isolation & purification*; D013154:Spleen / Q000378:metabolism*; D014982:Xenopus laevis / Q000378:metabolism
D004768:Enterotoxins; D008934:Mitogens; D012333:RNA, Messenger; D011061:Poly A; C026066:enterotoxin A, Staphylococcal; D012313:RNA; D007371:Interferon-gamma