Direct detection of RNA mediated by reverse transcriptase lacking RNAse H function

US5994079A · kind A · utility

Assignee

• Digene Corporation · US

Inventors

• Abel de la Rosa · Alpharetta, US
• Clayton D. Collier · Germantown, US

Key dates

• Filing date: Feb 6, 1998
• Grant date: Nov 30, 1999
• Expiry date: Feb 6, 2018

Classification

• Technology area (CPC C): Chemistry; Metallurgy
• CPC primary: C12Q1/6804
• WIPO field: Biotechnology
• WIPO sector: Chemistry

Abstract

Disclosed is a method of detecting RNA molecules of interest in which reverse transcription primers unique to the RNA molecule of interest are used for reverse transcribing the RNA with a reverse transcriptase lacking RNAse H function and the resulting RNA/DNA hybrid is detected with an antibody specific for RNA/DNA hybrids. This method can be used to detect the presence of one or many specific RNA molecules which may be present in a sample, including RNA from different organisms (such as viruses, bacteria, fungi, plants, and animals), or RNA indicative of an infection, a disease state, or predisposition to a disease in an animal. The specificity of detection is increased relative to current detection methods involving probe hybridization since the reverse transcription primers are shorter and less subject to non-specific hybridization. Specificity of the disclosed method can also be increased by using a thermostable reverse transcriptase and performing reverse transcription at a high temperature. The disclosed method can also be used to detect reverse transcriptase activity in a sample and to identify inhibitors of reverse transcriptase. Also disclosed is a method for sequencing t…