Biomaterial Database

Rapid DNA typing for class II HLA antigens: subtyping of DRw52-associated DRB1 alleles. 1993

C Horne, and P A Keown

Department of Medicine, Vancouver General Hospital, Canada.

Serologic typing for MHC class II antigens is incapable of identifying important subtypes for certain DRB1 alleles and occasionally leads to errors of assignment, particularly with the DR antigens associated with DRw52. To simplify DNA typing of DRw52-associated DRB1 alleles, we have developed a new rapid method using PCR-RFLP. The PCR-RFLP method is based on allele-specific amplification followed by digestion of PCR-amplified DNA with restriction enzymes. Group-specific amplification of the second exon of DR3, DR5, DR6 and DR8 was achieved using a 5' primer specific for the first hypervariable region sequence common to all alleles in this group and generic 3' primers. Human genomic DNA was amplified in a Perkin-Elmer Thermocycler. The presence of a 265 bp fragment was confirmed by agarose gel electrophoresis. Restriction enzyme digestion using Rsa I followed by polyacrylamide gel electrophoresis gave a pattern unique for some alleles and placed the remainder in subgroups. Digestion of the PCR product with one or two of the following enzymes (Asp 700, Hae II, Mnl I, Mbo II, Ksp I and Hph I) permitted the identification of 21 of the 22 alleles. DRB1*1103 and DRB1*1104 are not distinguished by this method and can be distinguished by SSOP or by using a specific 3' primer. For some heterozygous combinations, additional primers are used to provide full subtyping. This method provides a rapid and less costly alternative to PCR-SSOP for DRw52 subtyping in the smaller laboratory as only one amplification is required (two primers) for the majority of samples.(ABSTRACT TRUNCATED AT 250 WORDS)

UI	MeSH Term	Description	Entries
D008969	Molecular Sequence Data	Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories.	Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D012150	Polymorphism, Restriction Fragment Length	Variation occurring within a species in the presence or length of DNA fragment generated by a specific endonuclease at a specific site in the genome. Such variations are generated by mutations that create or abolish recognition sites for these enzymes or change the length of the fragment.	RFLP,Restriction Fragment Length Polymorphism,RFLPs,Restriction Fragment Length Polymorphisms
D005802	Genes, MHC Class II	Genetic loci in the vertebrate major histocompatibility complex that encode polymorphic products which control the immune response to specific antigens. The genes are found in the HLA-D region in humans and include H-2M, I-A, and I-E loci in mice.	Class II Genes,Genes, Class II,Genes, HLA Class II,MHC Class II Genes,Class II Gene,Gene, Class II
D006650	Histocompatibility Testing	Identification of the major histocompatibility antigens of transplant DONORS and potential recipients, usually by serological tests. Donor and recipient pairs should be of identical ABO blood group, and in addition should be matched as closely as possible for HISTOCOMPATIBILITY ANTIGENS in order to minimize the likelihood of allograft rejection. (King, Dictionary of Genetics, 4th ed)	Crossmatching, Tissue,HLA Typing,Tissue Typing,Crossmatchings, Tissue,HLA Typings,Histocompatibility Testings,Testing, Histocompatibility,Testings, Histocompatibility,Tissue Crossmatching,Tissue Crossmatchings,Tissue Typings,Typing, HLA,Typing, Tissue,Typings, HLA,Typings, Tissue
D006681	HLA-D Antigens	Human immune-response or Class II antigens found mainly, but not exclusively, on B-lymphocytes and produced from genes of the HLA-D locus. They are extremely polymorphic families of glycopeptides, each consisting of two chains, alpha and beta. This group of antigens includes the -DR, -DQ and -DP designations, of which HLA-DR is most studied; some of these glycoproteins are associated with certain diseases, possibly of immune etiology.	Antigens, HLA-D,Class II Human Antigens,HLA-Dw Antigens,Human Class II Antigens,Ia-Like Antigens, Human,Immune Response-Associated Antigens, Human,Immune-Associated Antigens, Human,Immune-Response Antigens, Human,HLA-D,HLA-Dw,Immune Response Associated Antigens, Human,Antigens, HLA D,Antigens, HLA-Dw,Antigens, Human Ia-Like,Antigens, Human Immune-Associated,Antigens, Human Immune-Response,HLA D Antigens,HLA Dw Antigens,Human Ia-Like Antigens,Human Immune-Associated Antigens,Human Immune-Response Antigens,Ia Like Antigens, Human,Immune Associated Antigens, Human,Immune Response Antigens, Human
D006684	HLA-DR Antigens	A subclass of HLA-D antigens that consist of alpha and beta chains. The inheritance of HLA-DR antigens differs from that of the HLA-DQ ANTIGENS and HLA-DP ANTIGENS.	HLA-DR,Antigens, HLA-DR,HLA DR Antigens
D006801	Humans	Members of the species Homo sapiens.	Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D000483	Alleles	Variant forms of the same gene, occupying the same locus on homologous CHROMOSOMES, and governing the variants in production of the same gene product.	Allelomorphs,Allele,Allelomorph
D000949	Histocompatibility Antigens Class II	Large, transmembrane, non-covalently linked glycoproteins (alpha and beta). Both chains can be polymorphic although there is more structural variation in the beta chains. The class II antigens in humans are called HLA-D ANTIGENS and are coded by a gene on chromosome 6. In mice, two genes named IA and IE on chromosome 17 code for the H-2 antigens. The antigens are found on B-lymphocytes, macrophages, epidermal cells, and sperm and are thought to mediate the competence of and cellular cooperation in the immune response. The term IA antigens used to refer only to the proteins encoded by the IA genes in the mouse, but is now used as a generic term for any class II histocompatibility antigen.	Antigens, Immune Response,Class II Antigens,Class II Histocompatibility Antigen,Class II Major Histocompatibility Antigen,Ia Antigens,Ia-Like Antigen,Ia-Like Antigens,Immune Response Antigens,Immune-Associated Antigens,Immune-Response-Associated Antigens,MHC Class II Molecule,MHC II Peptide,Class II Antigen,Class II Histocompatibility Antigens,Class II MHC Proteins,Class II Major Histocompatibility Antigens,Class II Major Histocompatibility Molecules,I-A Antigen,I-A-Antigen,IA Antigen,MHC Class II Molecules,MHC II Peptides,MHC-II Molecules,Antigen, Class II,Antigen, I-A,Antigen, IA,Antigen, Ia-Like,Antigens, Class II,Antigens, Ia,Antigens, Ia-Like,Antigens, Immune-Associated,Antigens, Immune-Response-Associated,I A Antigen,II Peptide, MHC,Ia Like Antigen,Ia Like Antigens,Immune Associated Antigens,Immune Response Associated Antigens,MHC II Molecules,Molecules, MHC-II,Peptide, MHC II,Peptides, MHC II
D001483	Base Sequence	The sequence of PURINES and PYRIMIDINES in nucleic acids and polynucleotides. It is also called nucleotide sequence.	DNA Sequence,Nucleotide Sequence,RNA Sequence,DNA Sequences,Base Sequences,Nucleotide Sequences,RNA Sequences,Sequence, Base,Sequence, DNA,Sequence, Nucleotide,Sequence, RNA,Sequences, Base,Sequences, DNA,Sequences, Nucleotide,Sequences, RNA