5-Bromo-2′-deoxyuridine

Technical Specifications

Product Description & Scientific Applications

5-Bromo-2′-deoxyuridine (BrdU) is a synthetic halogenated pyrimidine analogue and a marker of DNA synthesis: as a thymidine analogue, it is incorporated into newly synthesised DNA. BrdU-substituted DNA stains much more weakly with Hoechst dye and Giemsa than unsubstituted DNA. Bromine at the C5 position of pyrimidines, as in BrdU, has been used to attach a heavy atom covalently when derivatising DNA or RNA.

Cell proliferation assays. The BrdU incorporation assay is the most commonly used method for assessing DNA replication and is typically read out by antibody immunostaining of the incorporated label. Because the C5 bromide is also a cross-coupling handle, BrdU incorporated in cells can instead be labelled directly with fluorescent boronic acid probes by Suzuki–Miyaura coupling, without antibody immunostaining.

Sister chromatid exchange. Once BrdU has been incorporated over two cell cycles, Hoechst/Giemsa differential staining or anti-BrdU antibody detection reveals the newly synthesised DNA strand within each chromatid as harlequin chromosomes, which allows the exchanges to be analysed.

Cell-cycle kinetics. Flow-cytometric double staining gives bivariate BrdU/DNA distributions, from which tumour S-phase duration and other cell-kinetic parameters are calculated.

Replication timing. For genome-wide mapping, cells are pulse-labelled with BrdU, the early and late S-phase fractions are separated by flow sorting, and the labelled nascent DNA is immunoprecipitated and sequenced.

Nanopore sequencing. BrdU is detectable in DNA templates alongside other thymidine analogues, including CldU and IdU.

Radiosensitisation. Halogenated pyrimidines such as BrdU sensitise cells to ionising radiation only after they have been incorporated into cellular DNA in place of thymidine.

Photo-cross-linking. Site-specific substitution of BrdU into synthetic oligonucleotides greatly increases the yield of UV photo-cross-linking to bound proteins, which makes it a handle for mapping protein–DNA contacts.

Synthetic oligonucleotides and DNA fragments. As its phosphoramidite, 5-bromo-2′-deoxyuridine is incorporated into synthetic oligodeoxynucleotides for structure–activity comparisons with other modified pyrimidines in serum stability, duplex binding affinity and RNase H-directed cleavage. An enzymatic polymerase/ligase method places 5-bromo-2′-deoxyuridine at a predetermined site within double-stranded DNA fragments, enabling studies of the sequence dependence of sensitiser-induced DNA damage.

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