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The preparation of poly (dT)-5'-transferrin conjugates and hybridisation studies with poly (dA)-tailed linearised pBR322 plasmid DNA. 1988

S Weiler, and M Ariatti, and A O Hawtrey
Department of Biochemistry, University of Durban-Westville, South Africa.

The formation of transferrin-DNA complexes intended for ligand-directed transfection studies has been achieved through a hybridisation technique involving complementary homodeoxypolynucleotide chains attached to the participating protein and DNA species. Oligothymidylate residues (pT)n obtained by dicyclohexylcarbodiimide (CDI) polymerisation of thymidine-5'-monophosphate (5'-TMP) were activated to the 5'-imidazolides which on incubation with transferrin yielded the 5'linked phosphoramidates (pT)n-5'-transferrin. Homopolymeric chain extension of (pT)5-5'-transferrin by terminal transferase and dTTP at 30 degrees for 30 min yielded (pT) 300-5'-transferrin. Cleavage of the phosphoramide link in the polymer modified transferrin at 37 degrees was pronounced after 30 min although at 25 degrees hydrolysis was less than 5% after 4 hr. Poly(dT)-5'-transferrin readily hybridised with [3H]poly(dA)-tailed Pst 1 linearised pBR322 DNA. Resultant complexes were demonstrated by nitrocellulose filter binding and immunoprecipitation with anti-transferrin antibody. In contrast with poly(dT)-5'-transferrin, poly(dT)-5'-transferrin-poly(dA)-tailed pBR322 DNA complexes were stable at 37 degrees suggesting that annealing is followed by further stabilising interactions between the DNA and protein components.

UI MeSH Term Description Entries
D009693 Nucleic Acid Hybridization Widely used technique which exploits the ability of complementary sequences in single-stranded DNAs or RNAs to pair with each other to form a double helix. Hybridization can take place between two complimentary DNA sequences, between a single-stranded DNA and a complementary RNA, or between two RNA sequences. The technique is used to detect and isolate specific sequences, measure homology, or define other characteristics of one or both strands. (Kendrew, Encyclopedia of Molecular Biology, 1994, p503) Genomic Hybridization,Acid Hybridization, Nucleic,Acid Hybridizations, Nucleic,Genomic Hybridizations,Hybridization, Genomic,Hybridization, Nucleic Acid,Hybridizations, Genomic,Hybridizations, Nucleic Acid,Nucleic Acid Hybridizations
D010957 Plasmids Extrachromosomal, usually CIRCULAR DNA molecules that are self-replicating and transferable from one organism to another. They are found in a variety of bacterial, archaeal, fungal, algal, and plant species. They are used in GENETIC ENGINEERING as CLONING VECTORS. Episomes,Episome,Plasmid
D011071 Poly T A group of thymine nucleotides in which the phosphate residues of each thymine nucleotide act as bridges in forming diester linkages between the deoxyribose moieties. Poly dT,Polythymidylic Acids,Thymine Polynucleotides,Polydeoxythymidylate,Acids, Polythymidylic,Polynucleotides, Thymine,dT, Poly
D011089 Polydeoxyribonucleotides A group of 13 or more deoxyribonucleotides in which the phosphate residues of each deoxyribonucleotide act as bridges in forming diester linkages between the deoxyribose moieties. Polydeoxyribonucleotide
D013696 Temperature The property of objects that determines the direction of heat flow when they are placed in direct thermal contact. The temperature is the energy of microscopic motions (vibrational and translational) of the particles of atoms. Temperatures
D014168 Transferrin An iron-binding beta1-globulin that is synthesized in the LIVER and secreted into the blood. It plays a central role in the transport of IRON throughout the circulation. A variety of transferrin isoforms exist in humans, including some that are considered markers for specific disease states. Siderophilin,Isotransferrin,Monoferric Transferrins,Serotransferrin,Transferrin B,Transferrin C,beta 2-Transferrin,beta-1 Metal-Binding Globulin,tau-Transferrin,Globulin, beta-1 Metal-Binding,Metal-Binding Globulin, beta-1,Transferrins, Monoferric,beta 1 Metal Binding Globulin,beta 2 Transferrin,tau Transferrin

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