Patent ID: 7384769
Filing Date: 2008-06-10
Classification: C12Q

Abstract:
1. A method for the quantitative detection of a target nucleic acid sequence of HHV-6 from a sample, which comprises the following steps: a) extraction of the nucleic acid from the sample with another nucleic acid (calibrator) previously added to the sample itself, said calibrator having the same sequence of the target nucleic acid, said calibrator having i) the same sequence of the target nucleic acid, apart from the region hybridizing to the probe, which is randomized with respect to the corresponding region of the target nucleic acid, maintaining the same nucleotide composition, and ii) a Tm equal to the target nucleic acid Tm +/−4° C., b) mixing the extracted target nucleic acid and calibrator with a forward primer which comprises SEQ ID NO:25, with a reverse primer which comprises SEQ ID NO:26, with a target nucleic acid probe bearing a reporter and a quencher, said target probe comprising SEQ ID NO:27, and a calibrator probe bearing a reporter and a quencher, said calibrator probe comprising SEQ ID NO:28, and annealing said forward primer and said reverse primer with a nucleic acid polymerase with 5′-3′ nuclease activity, in suitable conditions to carry out a polymerization reaction, and c) determination of the signal associated with the reporters released due to the 5′ polymerase nuclease activity.