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Poly(A) + RNA from Tetrahymena: Stimulation of Protein Synthesis in Vitro * †
Author(s) -
MYHAL M. LYNN,
HUFNAGEL LINDA A.
Publication year - 1979
Publication title -
the journal of protozoology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.067
H-Index - 77
eISSN - 1550-7408
pISSN - 0022-3921
DOI - 10.1111/j.1550-7408.1979.tb04218.x
Subject(s) - rna , tetrahymena , biochemistry , lysis , rna extraction , nuclease , ribosomal rna , proteinase k , tetrahymena pyriformis , biology , microbiology and biotechnology , gel electrophoresis , dna , chromatography , chemistry , gene
SYNOPSIS Cell‐free synthesis of high molecular weight polypeptides, programmed by RNA from Tetrahymena pyriformis strain W is reported, and methods for preparation of the RNA are described. The RNA was extracted by the SDS‐phenol‐chloroform‐isoamyl alcohol technic. The bulk of extracted RNA was ribosomal and on sucrose gradients peaked at ‐17S and 25S. After heat denaturation all the 25S RNA was converted to 17S. indicating the presence of hidden breaks, possibly the result of nuclease activity during extraction. Nevertheless, when poly(A)–RNA was collected using oligo‐(dT)‐cellulose column chromatography, it promoted a 15–fold increase in incorporation of [35S] methionine into TCA‐precipitable material. Slab‐gel electrophoresis and autoradiography of the product revealed 12 different major polypeptides, varying in weight from 28.000 to 65,000 Daltons. A method for preparation of translatable RNA from Tetrahymena will make possible the comparison of messenger RNAs associated with specific cell structures and with different developmental events.