Biotin-16-dUTP
[CAS# 136632-31-0]

Identification

• Classification: Biochemical >> Nucleoside drugs >> Deoxynucleotides and their analogues
• Name: Biotin-16-dUTP
• Synonyms: 5-(N-[N-Biotinyl-e-aminocaproyl-?-aminobutyryl]-3-aminoallyl)-2'-deoxyuridine 5'-triphosphate
• Molecular Formula: C₃₂H₅₂N₇O₁₈P₃S
• Molecular Weight: 947.78
• CAS Registry Number: 136632-31-0
• SMILES: CC1(C(=O)N2C(S1)C(C2CCCCC(=O)NCCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCNC(=O)Nc3cc(c(=O)n(c3)C)C)NC3C(C(C(O3)COP(=O)(O)OP(=O)(O)OP(=O)(O)O)O)O)C

Properties

• Solubility: water: miscible (Expl.)

Discovery and Applications

Biotin-16-dUTP is a chemically modified nucleotide in which a biotin moiety is covalently attached to the 16th carbon position of the deoxyuridine triphosphate (dUTP) molecule via a linker. This modification allows biotin-16-dUTP to be incorporated into DNA strands during enzymatic DNA synthesis processes such as PCR, nick translation, or in situ hybridization. The incorporation of biotinylated nucleotides enables subsequent detection or purification of labeled DNA through high-affinity interactions with avidin or streptavidin proteins, which bind biotin with exceptional specificity and strength.

The discovery of biotinylated nucleotides arose from the need for sensitive and specific methods to detect nucleic acids in molecular biology. Traditional methods for DNA labeling, such as radioactive isotopes, posed challenges related to safety, stability, and disposal. Biotin labeling provided a non-radioactive alternative with robust detection capabilities due to the strong biotin-avidin interaction. The introduction of biotin-16-dUTP represented a significant advancement, as it allowed direct incorporation of biotin into DNA during synthesis without substantially disrupting the polymerase activity or DNA structure.

Applications of biotin-16-dUTP span a wide range of molecular biology techniques. In situ hybridization techniques utilize biotin-16-dUTP to label probes that bind complementary DNA or RNA sequences in fixed cells or tissue sections, allowing visualization of gene expression or chromosomal localization by staining with streptavidin conjugates linked to enzymes or fluorophores. Similarly, biotin-16-dUTP is widely used in enzyme-mediated labeling methods, such as nick translation or random priming, to generate biotin-labeled DNA probes for blotting or microarray analysis.

Beyond detection, biotinylated DNA can be immobilized on streptavidin-coated surfaces, facilitating nucleic acid purification, capture, or immobilization for downstream applications. This property is particularly valuable in affinity-based assays, DNA-protein interaction studies, and next-generation sequencing library preparations where selective capture of DNA fragments is essential.

The synthesis of biotin-16-dUTP involves conjugating biotin to dUTP through a spacer arm that preserves the nucleotide’s recognition by DNA polymerases. The 16-atom linker provides sufficient flexibility and distance between the biotin moiety and the nucleotide base, minimizing steric hindrance during enzymatic incorporation. This design ensures that DNA polymerases can efficiently incorporate biotin-16-dUTP alongside natural nucleotides during DNA synthesis.

In conclusion, biotin-16-dUTP is a crucial reagent in molecular biology that enables versatile and sensitive labeling of DNA for detection, purification, and analytical applications. Its discovery provided a non-radioactive, specific, and efficient method for tagging nucleic acids, which has been widely adopted in research and diagnostic protocols.

References

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