[Quantitative analysis of individual RNA in preparations of poly(A)+-RNA mammalian cells].
A new method of simultaneous analysis of the relative abundance of the most abundant individual mRNA's in poly(A)(+)-RNA preparations is described. The method is based on the synthesis of short (10-20 nucleotides) cDNA products by reverse transcription of poly(A)(+)-RNA primed with 5'-labeled oligonucleotides of 9 nucleotide lengths. Three natural nucleotides and one terminator nucleotide are used as substrates for reverse transcriptase. The numbers, lengths and sequence of the oligonucleotides used as primers were chosen to provide more than a 90% probability that synthesis would be initiated from any individual RNA present in the poly(A)(+)-RNA, thus assuring comprehensive analysis of RNA with abundance higher than 0.01%. Each primer produces about 20-60 bands per track following polyacrylamide electrophoresis under denaturing conditions. A full set of 30 oligonucleotides used to analyze a poly(A)(+)-RNA preparation produces an electrophoretic pattern with information capacity similar to that obtained from high resolution 2-dimensional electrophoresis of protein. Using this method we show that the patterns of poly(A)(+)-RNA differ from tissue to tissue, from normal tissues to neoplastic tissue (human myoma of uterus) and during differentiation of a F9 embryonic carcinoma cell line.
D001483:Base Sequence; D002478:Cells, Cultured; D004247:DNA / Q000138:chemical synthesis; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008969:Molecular Sequence Data; D009841:Oligonucleotides; D011061:Poly A / Q000378:metabolism*; D012313:RNA / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis*
D009841:Oligonucleotides; D012333:RNA, Messenger; D011061:Poly A; D012313:RNA; D004247:DNA
Kolichestvennyĭ analiz individual'nykh RNK v preparatakh poli(A)+-RNK kletok mlekopitaiushchikh.