Abstract
DNA replication was investigated in cells with polytene chromosomes. The cells were obtained from the salivary glands of the dipteran Chironomus tentans. Polytene chromosomes are characterized by a specific and constant band — interband structure formed by the lateral association of homologous chromatids side by side. — The salivary gland DNA was labelled by injection of radioactive precursor into the living animal, extracted with a neutral nondenaturing buffer at 25° C and finally characterized by agarose gel electrophoresis. Radioactive DNA pulse-labelled for 30–60 min was released from the polytene chromosomes during cell lysis in the form of double-stranded fragments. The fragments, which show a heterogeneous appearance in gel electrophoresis, are probably produced in the living cell by the joining of several Okazaki fragments. The release of the fragments from the polytene chromosome is prevented by lysis at 0° C instead of 25° C. The size of the double-stranded fragments range between 3.75–6×106 D. Moreover, after a time-lag the fragments are joined together to produce a high-molecular weight DNA. The existence of these nascent DNA fragments is discussed in relation to an earlier proposal that each band in the polytene chromosome may function as a separate replication unit.
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References
Alberts, B., Sternglanz, R.: Recent excitement in the DNA replication. Nature (Lond.) 269, 655–661 (1977)
Beermann, W.: Cytologische Analyse eines Camptochironomus-Artbastards. I. Kreuzungsergebnisse und die Evolution des Karyotypus. Chromosoma (Berl.) 7, 198–259 (1955)
Beermann, W.: Chromosomes and genes. In: Developmental studies on giant chromosomes (W. Beermann, ed.), pp. 1–33. Berlin-Heidelberg-New York: Springer 1972
Blumenthal, A.B., Kriegstein, H.J., Hogness, D.S.: The units of DNA replication in Drosophila melanogaster chromosomes. Cold Spring Harbor Symp. quant. Biol. 38, 205–223 (1973)
Callan, H.G.: Replication of the DNA in the chromosomes of eukaryotes. Phil. Trans. roy. Soc. (Lond.) B 181, 19–41 (1972)
Daneholt, B., Edström, J.-E.: The content of deoxyribonucleic acid in individual polytene chromosomes of Chironomus tentans. Cytogenetics 6, 350–356 (1967)
Darrow, J., Clever, U.: Chromosome activity and cell function in polytenic cells. III. Growth and replication. Develop. Biol. 21, 331–348 (1970)
Edström, J.-E.: Microextraction and microelectrophoresis for determination and analysis of nucleic acids in isolated cellular units. In: Methods in cell physiology (I.D.M. Prescott, ed.), pp. 417–447. New York: Academic Press 1964
Edström, J.-E., Lönn, U.: Cytoplasmic zone analysis. RNA flow studied by micromanipulation. J. Cell Biol. 70, 562–672 (1976)
Guy, A., Taylor, J.H.: Actinomycin D inhibits initiation of DNA replication in mammalian cells. Proc. nat. Acad. Sci. (Wash.) 75, 6088–6092 (1978)
Hägele, K.: Prolongation of replication times after doublings of the DNA content of polytene chromosome bands of Chironomus. Chromosoma (Berl.) 55, 253–258 (1976)
Hand, R.: Eukaryotic DNA: Organization of the genome for replication. Cell 15, 317–325 (1978)
Helling, R., Goodman, H., Boyer, H.: Analysis of endonuclease EcoRI fragments of DNA from lambdoid bacteriophages and other viruses by agarose-gel electrophoresis. J. Virol. 14, 1235–1244 (1974)
Hollenberg, C.P.: Proportionate representation of rDNA and Balbiani ring DNA in polytene chromosomes of Chironomus tentans. Chromosoma (Berl.) 57, 185–197 (1976)
Lambert, B.: Repeated nucleotide sequences in polytene chromosomes. Thesis. Karolinska Institute!, Stockholm, Sweden (1972)
Lönn, U.: Delayed flow through cytoplasm of newly synthesized Balbiani ring 75 S RNA. Cell 13, 727–733 (1978)
Lönn, U., Edström, J.-E.: Movements and associations of ribosomal subunits in a secretory cell during growth inhibition by starvation. J. Cell Biol. 73, 696–704 (1977)
Pelling, C.: A replicative and synthetic chromosomal unit — the modern concept of the chromomere. Proc. roy. Soc. (Lond.) B 164, 279–289 (1966)
Rudkin, G.T.: Replication in polytene chromosomes. In Developmental Studies on giant chromosomes (W. Beermann, ed.), pp. 58–86. Berlin-Heidelberg-New York: Springer 1972
Taylor, J.H.: Replication of DNA in mammalian chromosomes: Isolation of replicating segments. Proc. nat. Acad. Sci. (Wash.) 70, 1083–1087 (1973)
Thomas, M., Davis, R.: Studies on the cleavage of Bacteriophage lambda DNA with EcoRI restriction endonuclease. J. molec. Biol. 91, 315–328 (1975)
Wells, R., Royer, H.-D., Hollenberg, C.: Non Xenopus-like DNA sequence organization in Chironomus tentans genome. Molec. gen. Genet. 147, 45–51 (1976)
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Lönn, U. Isolation and partial characterization of replication intermediates in Chironomus polytene chromosomes. Chromosoma 77, 29–40 (1980). https://doi.org/10.1007/BF00292039
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DOI: https://doi.org/10.1007/BF00292039