Heteroduplex tracking assay (HTA) for genotyping HCV
AssigneeChiron Corporation
InventorAmy J. Weiner
Life of this patent
Estimating the term…
Abstract
A heteroduplex tracking assay (HTA), a hybridization based method of determining the genetic relationship between two or more viral genomes, for genotyping HCV is disclosed. The HTA for genotyping HCV was developed using single stranded probes derived form the carboxyl terminus of core and part of the E1 for HCV subtypes 1a, 1b, 2a, 2b, and 3a. HTA is more accurate than RFLP for sub-typing HCV and has potential for identifying new variants and is useful for epidemiological studies.
Claims23
An oligonucleotide consisting of the sequence of Seq ID No. 1.
An oligonucleotide consisting of the sequence of Seq ID No. 2.
An oligonucleotide consisting of the sequence of Seq ID No. 3.
An oligonucleotide consisting of the sequence of Seq ID No. 4.
An oligonucleotide consisting of the sequence of Seq ID No. 5.
An oligonucleotide consisting of the sequence of Seq ID No. 6.
An oligonucleotide consisting of the sequence of Seq ID No. 7.
An oligonucleotide consisting of the sequence of Seq ID No. 8.
An oligonucleotide consisting of the sequence of Seq ID No. 9.
A pair of PCR primers wherein the sense primer consists of Seq ID NO. 1 and the antisense primer is selected from the group consisting of Seq ID NO 2, Seq ID NO. 3, Seq ID NO. 4 and Seq ID NO. 5.
A pair of PCR primers wherein the antisense primer consists of Seq ID NO. 6 and the sense primer is selected from the group consisting of Seq ID NO 7, Seq ID NO. 8, and Seq ID NO. 9.
A method of determining the HCV genotype of an HCV strain, said method comprising the steps of:
- (a) subjecting said HCV strain to one or more stages of PCR, wherein the one or more stages of PCR utilizes a sense probe from the core region of the HCV genome and an antisense probe from the E1 region of the HCV genome, or a sense probe from the E1 region of the HCV genome and an antisense probe from the core region of the HCV genome;
- (b) forming a heteroduplex by denaturing and reannealing mixtures of the amplified product obtained in step (a) with DNA or RNA fragments of a known HCV genotype, said fragments comprising a region of the HCV genome between the core and E1 regions;
- (c) comparing the mobility of said heteroduplex on a system that separates by size with the mobility of a homoduplex of the DNA or RNA fragments of known genotype to determine the genotype of the HCV strain.
Dependencies are inferred from the claim text.