DNA sequence-based HLA typing method
US5972604A · kind A · utility
Assignee
Inventors
Key dates
| Filing date | Aug 23, 1996 |
| Grant date | Oct 26, 1999 |
| Priority date | — |
| Expiry date | Aug 23, 2016 |
Classification
- Technology area (CPC C)Chemistry; Metallurgy
- CPC primaryC12Q2600/156
- WIPO fieldBiotechnology
- WIPO sectorChemistry
Abstract
The present invention provides a process for determining genotypes in highly polymorphic systems by polymerase chain reaction amplification of CDNA or genomic DNA and direct sequencing polymerase chain reaction products using oligonucleotide primers. More specifically, Class II and Class I HLA genotypes can be unambiguously determined in any subject in 16-24 hours by direct sequencing of DRB, DQB, DQA, DPB, DPA, HLA-A, HLA-B and HLA-C- transcripts enzymatically amplified using a limited number of non-allele-specific oligonucleotides. Total cellular RNA from peripheral blood mononuclear cells is reverse transcribed using antisense primers, specific for different locus (DQB, DQA, DPA or DPB) or group of loci (DRB1-5, or HLA-A and HLA-B and HLA-C). The synthesized cDNA molecules are then enzymatically amplified using different combinations of oligonucleotides for each locus and directly sequenced with Taq polymerase using an internal oligonucleotide. The sequenced genes are then analyzed.
Source: USPTO / EPO open patent data. Objective bibliographic and citation counts.