Repetitive and non-repetitive sequence in sea urchin heterogeneous nuclear RNA

Abstract

Heterogeneous nuclear RNA was extracted from sea urchin gastrulae following a ten-minute pulse with [^3H]uridine. Hybridization reactions with excess sheared DNA were carried out using several methods. The fraction of the Hn[^3H]RNA in hybrids was determined by RNAase resistance and hydroxyapatite chromatography in a urea-phosphate medium. After incubation to a DNA C_0t value of 40, 20 to 25% of the Hn[^3H]RNA binds to hydroxyapatite. Approximately one third of these bound molecules are in repetitive DNA-RNA hybrids. The size of the bound RNA molecules measured after hybridization averages 1100 nucleotides. Up to 70% of the Hn[^3H]RNA forms hybrids at high DNA/RNA ratios (130,000/1) when reactions are carried out to high C0t values. Studies of the effect of increased DNA/RNA ratio on extent of hybridization at high DNA C_0t values show that at least two frequency classes of single copy transcript are present in the Hn[3H]RNA population. Both repetitive and single copy DNA transcripts are present within individual Hn[^3H]RNA molecules. Calculations indicate that at least 25% and possibly all of the Hn[^3H]RNA molecules contain repetitive sequence elements interspersed with non-repetitive sequences.

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