Non-enzymatic in vitro DNA labeling and label immunoquantification.
In the present work, a label immunoquantification procedure was developed in order to determine the number of markers introduced into DNA. A non-enzymatic, in vitro labeling method for introducing the p-bromobenzoyl radical (label), through transamination and acylation reactions of the cytidine nucleotides in calf thymus DNA, was carried out. Three spacer arms with different lengths were used for separating the label from the nucleotide and three labeled DNA were obtained. Anti-p-bromobenzoyl chicken IgY polyclonal antibodies were obtained. The antibodies detected the label, into three-labeled DNA, with different sensitivities, in relation to spacer arm length used. About 3-11 labels per 4 x 10(6) bases into thermally denatured DNA were immunoquantified.
Clarke
David Higginson
DH
Molecular Biology Department, Neurosciences Center, Ave. 25, No. 15202, Playa, Havana A.P. 6990, Cuba.
Pérez-Bello
Dannelys
D
Riverón-Rojas
Ana María
AM
Jiménez-Alemán
Guillermo
G
Acosta
Mirta Zayas
MZ
Perera
Rafaela Pérez
RP
Carrazana
Marquiza Sablón
MS
Miranda
Nelson Díaz
ND
Rodríguez-Tanty
C
C
Gutiérrez Calzado
Esteban
E
D000818:Animals; D002645:Chickens; D004247:DNA / Q000032:analysis* / Q000737:chemistry*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D004798:Enzymes / Q000737:chemistry; D015151:Immunoblotting / Q000379:methods*; D007136:Immunoglobulins / Q000032:analysis* / Q000737:chemistry*; D015335:Molecular Probes / Q000032:analysis / Q000737:chemistry; D013194:Staining and Labeling / Q000379:methods*
D004798:Enzymes; C016512:IgY; D007136:Immunoglobulins; D015335:Molecular Probes; D004247:DNA; C427203:calf thymus DNA