Open AccessIsolation and Characterization of Xenopus laevis Albumin mRNA
Author(s) -
DIMITRIADIS Giorgos J.
Publication year - 1981
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1981.tb06394.x
Subject(s) - albumin , microbiology and biotechnology , messenger rna , biochemistry , bovine serum albumin , gel electrophoresis , centrifugation , biology , rna , serum albumin , xenopus , chromatography , agarose gel electrophoresis , agarose , chemistry , dna , gene
The messenger RNA for albumin was isolated from the liver of male frogs. Purification was achieved using oligo(dT)‐cellulose chromatography and sucrose gradient centrifugation under denaturing conditions. As judged by translational activity in a cell‐free protein‐synthesizing system derived from rabbit reticulocytes, albumin mRNA was enriched 259‐fold as compared to the total ribonucleic acid of the liver cells. Purified albumin mRNA migrated after sucrose gradient centrifugation as a single symmetrical peak of approximately 17 S and also moved as a single band following denaturing agarose gel electrophoresis. Albumin mRNA possesses properties compatible with the presence of a poly(adenylic acid) sequence. Translation in vitro yielded a product which is immunoprecipitable with anti‐(frog albumin) and which showed a single radioactive peak having a molecular weight of about 74000 in sodium dodecylsulfate/polyacrylamide gel electrophoresis. Complementary DNA was synthesized using reverse transcriptase and, as a template, purified albumin mRNA. Following hybridization under conditions of excess RNA, the r 0 t 1/2 of albumin mRNA was found to be 1.8 × 10 −3 mol · s · 1 −1 . This result also confirmed that albumin mRNA had been isolated in a highly purified form.