Method for the amplification based on transcription of a nucleic acid sequence of HBV DNA target based on double filament of the HBV optionally present in a sample, oligonucleotide, and.Test kit suitable for performing amplification and detection based on transcription of the HBV DNA

摘要

Method for the amplification based on transcription of a nucleic acid sequence of HBV DNA of HBV target on the basis of the optionally present in a sample, the oligonucleotide.Set of oligonucleotide primers suitable for use in nucleic acid amplification of HBV, and.Test kit suitable for performing the amplification and detection of DNA based on transcription of the HBV.The present invention provides a method to an amplification based on transcription of a nucleic acid sequence of HBV target on the DNA of HBV optionally present in a sample.It comprises the steps of incubate the sample suspected to contain HBV.In a buffer amplifier with one or more restriction enzymes capable of cleaving the DNA of HBV in a restriction site selected.This restriction enzyme, creating a tip 3 39 set (s) from said filament (s) of the HBV DNA, an initiator promoter.The said initiator promoter having a region 5 39 which comprises the sequence of a promoter recognized by the RNA polymerase on DNA and a region 3 39 complementary to the extremity 3 39 def...Ined from the filament of a second or reverse primer DNA, having the opposite polarity of the initiator promoter and covers the tip 5 39 of the target sequence.And in the case of ssDNA of HBV as the target sequence, an initiator of the restriction.- keep the mixture of reaction thus created under the appropriate conditions for a sufficient amount of time for digestion by the restriction enzyme occurs.Submit the sample thus obtained to a heat treatment at a temperature and time sufficient to inactivate the enzyme restriction and / or to become at least partially a double filament in filameNT only, add the following reagents to the sample: an enzyme having activity of DNA polymerase dependent on RNA, an enzyme having activity of DNA dependent DNA polymerase.An enzyme having activity of RNase H,An enzyme having the activity of RNA polymerase and keep the reaction mixture thus created under the appropriate conditions for a sufficient amount of time for the amplification occurs.