We have investigated the locations of the poly(A), the mRNA-specific sequences and the RNA sequences that are eventually cleaved from the 1860 nucleotide precursor of mouse beta-globin mRNA. Biochemical and electron microscopic data demonstrate that there are two intervening RNA moieties in the precursor which separate the beta-globin mRNA sequences into three portions containing 480, 205 and 155 nucleotides. One of the two intervening RNA moieties contains 780 nucleotides. The size of the smaller intervening RNA has not been determined precisely, but it is 125 nucleotides or less. The largest mRNA-specific fragment is derived from the 3' terminus of the precursor, and contains the 3' terminal poly (A) and 330 mRNA-specific transcribed nucleotides. At least one, and probably both, intervening RNAs occur within the coding portion of the mRNA sequences. The larger of the intervening RNAs is located next to the 480 nucleotide mRNA-specific fragment, and the smaller intervening RNA is located between the 205 and 155 nucleotide mRNA-specific fragments. These experiments are consistent with the notion that the intervening sequences in the DNA of mouse beta-globin genes are transcribed into the mRNA precursor and are excised from the RNA by post-transcriptional events.