A one-tube nucleic acid extraction and amplification (Ertpacr) method for detecting RNA viruses

摘要

Extraction Reverse Transcription Polymerase Chain Reaction (ERTPCR), a rapid and simple method which allows detection of RNA viruses, is described. Unlike conventional Reverse Transcription Polymerase Chain Reaction(RTPCR), which requires extraction of viral nucleic acid prior to reverse transcription and DNA amplification, ERTPCR allows these procedures to be accomplished in one step in the same reaction tube. This is facilitated through heat extraction of viral RNA, and the use of a thermostable enzyme which can perform reverse transcription and DNA amplification in the same buffer system. The technique is at least as sensitive as a standard RTPCR. Its use in detecting rotavirus in feces, and hepatitis A virus in infected tissue cultures, is demonstrated. A silica spin column was used to remove RTPCR inhibitors from these samples.