[Inclusion of biotinylated analogs of dUTP and dCTP in DNA by DNA-polymerases. Cloning DNA fragments, containing biotinylated deoxyribouridine in E. coli].
The synthesis of a biotinylated derivative of dCTP, viz. N4-[(N-biotinyl)-4-amino-butoxyl]-2'-deoxycytidine 5'-triphosphate (I), is described. DNA polymerase I (Klenow fragment) incorporates (I) in DNA chains instead of thymidine, although with a lower efficiency than previously described biotinylated dUTP derivative (II), whereas highly purified DNA polymerase alpha from human placenta uses as substrate derivative (II) but not (I). A DNA fragment bearing biotin residues in one of strands was synthesized with the use of DNA polymerase alpha and dUTP derivative (II); its cloning in the plasmid vector pBR322 revealed that the DNA nucleotide sequence remained intact.
Oshevskiĭ
S I
SI
Kumarev
V P
VP
Grachev
S A
SA
D001483:Base Sequence; D001710:Biotin; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics*; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D003845:Deoxycytosine Nucleotides / Q000378:metabolism*; D003856:Deoxyuracil Nucleotides / Q000378:metabolism*; D004587:Electrophoresis, Agar Gel; D004926:Escherichia coli / Q000235:genetics*; D005260:Female; D005798:Genes, Bacterial; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D015345:Oligonucleotide Probes; D010920:Placenta / Q000378:metabolism; D011247:Pregnancy; D014169:Transformation, Bacterial
D003845:Deoxycytosine Nucleotides; D003856:Deoxyuracil Nucleotides; D015345:Oligonucleotide Probes; D001710:Biotin; D004247:DNA; D004259:DNA-Directed DNA Polymerase
Vkliuchenie v DNK biotinilirovannykh analogov dUTP i dCTP DNK- polimerazami. Klonirovanie v E. coli fragmenta DNA, soderzhashchego biotinilirovannyĭ dezoksiribouridin.