Sister chromatid differentiation with biotin-dUTP.

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.