[Structure of nuclear pre-mRNA. XIII. Hybridization properties of triphosphorylated 5'-end fragments]

Triphosphorylated 5'-end fragments about 100 nucleotides long were prepared from purified nuclear pre-mRNA using a modified hydroxyapatite method. These fragments as well as fragments of total pre-mRNA were polyadenylated by ATP:RNA adenyl-transferase and used as templates for the synthesis of [32P]cDNA by reverse transcriptase in the presence of an oligo(dT)-primer, cDNA transcribed from total fragments of pre-mRNA and from 5'-end fragments (5'-cDNA) were hybridized with excess of nuclear pre-mRNA. The extent of hybridization was 65-70 and 80-85% in different experiments. 18% of total cDNA and 35% of 5'-cDNA hybridized with mRNA from polysomes. A high homology between mRNA and triphosphorylated 5'-ends of pre-mRNA may be explained in the terms of splicing. The sequences adjacent to the triphosphorylated 5'-ends of pre-mRNA represent a specific class with complexity about 2.10(5) nucleotides. Less than 30% of 5'-cDNA hybridized with intermediately repetitive DNA, while the main portion hybridized with unique DNA sequences. About 15% of 5'-cDNA contain oligo (dA) sequences, originated from oligo (U) in the pre-mRNA.