Method of preparing and applying single stranded DNA probes to double stranded target DNAs in situ
US5028525A · kind A · utility
Assignee
Inventors
Key dates
| Filing date | Jun 26, 1990 |
| Grant date | Jul 2, 1991 |
| Priority date | — |
| Expiry date | Jun 26, 2010 |
Classification
- Technology area (CPC Y)Emerging Cross-Sectional Technologies
- CPC primaryY10T436/143333
- WIPO fieldBiotechnology
- WIPO sectorChemistry
Abstract
A method is provided for producing single stranded non-self-complementary nucleic acid probes, and for treating target DNA for use therewith. Probe is constructed by treating DNA with a restriction enzyme and an exonuclease to form template/primers for a DNA polymerase. The digested strand is resynthesized in the presence of labeled nucleoside triphosphate precursor. Labeled single stranded fragments are separated from the resynthesized fragments to form the probe. Target DNA is treated with the same restriction enzyme used to construct the probe, and is treated with an exonuclease before application of the probe. The method significantly increases the efficiency and specificity of hybridization mixtures by increasing effective probe concentration by eliminating self-hybridization between both probe and target DNAs, and by reducing the amount of target DNA available for mismatched hybridizations.
Source: USPTO / EPO open patent data. Objective bibliographic and citation counts.