Labelling of RNA and DNA moieties of mammalian embryos in-vivo by 14C-U-glucose and 32P-orthophosphate
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Uniformly labelled 14C-glucose and 32P-orthophosphate were applied intravenously to pregnant rats on day 12 of gestation. DNA and RNA were found to be labelled by both of these precursors.
Highly purified RNA and DNA were isolated from the embryos and hydrolyzed to mononucleotides and bases, the four nucleotides or bases were separated and the specific activities determined. According to the alkaline hydrolysis, a nearest-neighbour analysis (2′–3′-nucleotides) is achieved in the case of the RNA nucleotides whereas the enzymic hydrolysis of the DNA allows the preparation of 5′-nucleotides.
The ribosomal RNA bases were found to be labelled rather equally (≃1.4dpm/nmol) except for Uracil where the specific activity showed a 3 times higher value (≃4 dpm/nmol).
The base moieties of the DNA show an almost equal rate of incorporation of 14C-glucose fragments. The 32P-radioactivity found in DNA nucleotides is in the range of the corresponding 14C-data except for that of TMP. In this case a higher rate of 32P was found compared to the 14C-radioactivity.
Key wordsMethod 14C-Glucose 32P-Orthophosphate Rat Embryos DNA, RNA
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