Open AccessMolecular‐Weight Determination of Animal‐Cell RNA by Electrophoresis in Formamide under Fully Denaturing Conditions on Exponential Polyacrylamide Gels
Author(s) -
SPOHR Georges,
MIRAULT MarcEdouard,
IMAIZUMI Tereza,
SCHERRER Klaus
Publication year - 1976
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1976.tb10163.x
Subject(s) - rna , formamide , molecular mass , polyacrylamide gel electrophoresis , microbiology and biotechnology , ribosomal rna , gel electrophoresis , transfer rna , electrophoresis , dna , biology , hela , polyacrylamide , denaturation (fissile materials) , 5s ribosomal rna , chemistry , biochemistry , cell , gene , genetics , enzyme , nuclear chemistry
A method for electrophoretic analysis of RNA under fully denaturing conditions on exponential gradient polyacrylamide gels is described. Full denaturation, and strand separation of DNA · RNA hybrids and double‐stranded RNA is obtained in dry formamide only if electrophoresis is carried out at 45° and 55 °C, respectively. In such conditions, the effects of secondary structure of RNA, important in aqueous medium, are suppressed and a linear correlation is obtained between the logarithm of the molecular weight of an RNA and its final position in the gel over the entire molecular weight range of 10 4 –10 7 . Based on absolute molecular weight standards, obtained from sequenced rRNA of Escherichia coli and tRNA and extrapolating to higher molecular weights, the size of animal cell RNA was reexamined. Precursor rRNA (45 S) from HeLa cells migrates according to a molecular weight of 4.1 × 10 6 . Nascent precursor mRNA has molecular weights of up to 5 × 10 6 in the case of duck erythroblasts and of up to 10 7 in HeLa cells. This seems to represent the largest size of non‐viral animal‐cell RNA molecules.