Nick translation of HeLa cell nuclei as a probe for locating DNase I-sensitive nucleosomes.
The technique of nick translation of nuclei (Levitt, A., Axel, R., and Cedar, H. (1979) Dev. Biol. 69, 496-505) has been used in HeLa cells to label DNase I-sensitive regions. Micrococcal nuclease digestion of the nick translated nuclei was followed by a low ionic strength gel electrophoresis system which separates different types of mononucleosomes. The major label was observed in the vicinity of high mobility group protein containing mononucleosomes. However, further analysis revealed that the particle does not sediment in the position of mononucleosomes on a sucrose gradient. Two alternative explanations are discussed as the possible source of this particle. It is either a high mobility group protein containing nucleosome in some unfolded conformation or the labeled particle originates from discrete DNA fragments, wrapped around some nonhistone proteins, located in a highly DNase I-sensitive region, which is resistant to micrococcal nuclease digestion.
D002467:Cell Nucleus / Q000378:metabolism*; D004273:DNA, Neoplasm / Q000302:isolation & purification; D003850:Deoxyribonuclease I; D004706:Endodeoxyribonucleases / Q000378:metabolism*; D006367:HeLa Cells / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008836:Micrococcal Nuclease; D008970:Molecular Weight; D009363:Neoplasm Proteins / Q000302:isolation & purification; D009698:Nucleoproteins / Q000302:isolation & purification; D009707:Nucleosomes / Q000648:ultrastructure*; D014176:Protein Biosynthesis
D004273:DNA, Neoplasm; D009363:Neoplasm Proteins; D009698:Nucleoproteins; D009707:Nucleosomes; D004706:Endodeoxyribonucleases; D003850:Deoxyribonuclease I; D008836:Micrococcal Nuclease