A rapid and simple method for labeling short DNA fragments using Taq polymerase.
Biotechniques
; 12(3): 350-4, 1992 Mar.
Article
em En
| MEDLINE
| ID: mdl-1571140
ABSTRACT
This report describes a new method for labeling PCR-generated short length (60-120 bp) double-stranded DNA fragments for use as hybridization probes. The method utilizes gene-specific primers identical to those for PCR generation of non-radioactive DNA fragments. Radioactive probes are synthesized by Taq DNA polymerase without using PCR. Single-stranded (sense or antisense) and double-stranded probes can be individually prepared by selection of the appropriate primers. The labeling reaction reached maximum incorporation within 30 min with mean specific activities of 1.05 x 10(9) dpm/microgram (antisense single-stranded), and 1.62 x 10(9) dpm/microgram (double-stranded) were obtained using templates 69-117 of nucleotides. This method offers a simple and rapid means of generating antisense probes for Northern blot analyses and double-stranded probes for Southern blot analyses that provide highly intense signals with low background.
Buscar no Google
Base de dados:
MEDLINE
Assunto principal:
DNA
/
DNA Polimerase Dirigida por DNA
Tipo de estudo:
Diagnostic_studies
/
Evaluation_studies
Limite:
Animals
/
Pregnancy
Idioma:
En
Ano de publicação:
1992
Tipo de documento:
Article