Abstract
The method of choice to differentiate sister chromatids is to incorporate BrdU in replicating DNA. The disadvantage of BrdU is that its spontaneous or induced radicalization may itself lead to sister chromatid exchanges. Biotin-labelled dUTP is a widely used thymidine analogon for labelling isolated DNA. Its chemical structure suggests that, in contrast to BrdU, it does not give rise to radical formation. We electroporated proliferating Chinese hamster ovary (CHO) cells in the presence of biotin-dUTP which was subsequently detected in metaphase cells with TRITC-conjugated avidin. Microscopic analysis of second mitoses after labelling revealed a clear differential staining of sister chromatids. Thus substitution of thymidine with biotin-dUTP is another method to analyse SCE.
Similar content being viewed by others
References
Banfalvi G, Wiegant J, Sarkar N, van Duijn P (1989) Immunofluorescent visualization of DNA replication sites within nuclei of Chinese hamster ovary cells. Histochemistry 93: 81-86.
Folle GA, Boccardo E, Obe G (1997) Localization of chromosome breakpoints induced by DNase I in Chinese hamster ovary (CHO) cells. Chromosoma 106: 391-399.
Hiriyanna KT, Varkey J, Beer M, Benbow RM (1988) Electron microscopic visualization of sites of nascent DNA synthesis by streptavidin-gold binding to biotinylated nucleotides incorporated in vivo. J Cell Biol 107: 33-44.
Huijzer JC, Smerdon MJ (1992) Characterization of biotinylated repair regions in reversibly permeabilized human fibroblasts. Biochemistry 31: 5077-5084.
Hunting DJ, Dresler SL, de Murcia G (1985) Incorporation of biotin-labelled deoxyuridine triphosphate into DNA during excision repair and electron miroscopic visualization of repair patches. Biochemistry 24: 5729-5734.
Johannes C, Obe G (1991) Induction of chromosomal aberrations with the restriction endonuclease Alul in Chinese hamster ovary cells: comparison of different treatment methods. Int J Radiat Biol 59: 1379-1393.
Morris SM (1991) The genetic toxicology of 5-bromodeoxyuridine in mammalian cells. Mutat Res 258: 161-188.
Natarajan AT, Rotteveel AHM, van Pieterson J, Schliermann MG (1986) Influence of incorporated 5-bromodeoxyuridine on the frequencies of spontaneous and induced sister-chromatids exchanges, detected by immunological methods. Mutat Res 163: 51-55.
Perry P, Wolff S (1974) New Giemsa method for the differential staining of sister chromatids. Nature 251: 156-158.
Richer C-L, Drouin R (1990) Dynamic banding for high-resolution analysis of chromosomes and assignment of DNA replication times. In: G. Obe, ed. Advances in Mutagenesis Research, Vol 2. Berlin: Springer-Verlag, pp 55-94.
Schunck C (1997) Zur Entstehung von Schwesterchromatidenaustauschen und Chromosomenaberrationen. Experimente mit verschiedenen DNS-Doppelstrangbruch-induzierenden Agenzien. Dissertation, Universität Essen. Berlin: Logos-Verlag.
Sutou S (1981) Spontaneous sister-chromatid exchanges in Chinese hamster cells in vivo and in vitro. Mutat Res 82: 331-341.
Taylor JH (1958) Sister chromatid exchanges in tritium-labelled chromosomes. Genetics 43: 515-529.
Wolff S (1982) Sister Chromatid Exchange. New York: John Wiley & Sons.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Bruckmann, E., Wojcik, A. & Obe, G. Sister Chromatid Differentiation With Biotin-dUTP. Chromosome Res 7, 185–189 (1999). https://doi.org/10.1023/A:1009295131877
Issue Date:
DOI: https://doi.org/10.1023/A:1009295131877