HLA matching of unrelated bone marrow transplant pairs: direct sequencing of in vitro amplified HLA-DRB1 and -DQB1 genes using magnetic beads as solid support.

Sequencing of HLA genes can offer complete information on the HLA class II genes relevant for the outcome of bone marrow transplantation (BMT). Genomic HLA matching of unrelated BMT patient/donor pairs is often based on PCR-SSO typing of HLA class II alleles. Typing a small number of samples by this approach is both expensive and time-consuming, due to the large number of SSO probes required to perform a complete class II typing. Moreover, only polymorphisms explicitly tested for will be found. We now provide the first report of the use of direct sequencing of HLA-DRB1 and -DQB1 genes, using PCR-amplified genomic DNA attached to magnetic beads, for clinical routine HLA matching. Sequencing ladders obtained by this procedure are easily readable, the patterns can be interpreted in HLA homozygous as well as heterozygous individuals, and sequence differences or similarities between the BMT donor and recipient can be directly identified. This genomic typing method is informative, relatively fast and therefore well-suited for the small number of samples usually analyzed in matching of BMT pairs. Furthermore, this technique has the potential for automation.

[1]  J. Bodmer,et al.  HLA Class II nucleotide sequences, 1992. , 1993, Immunobiology.

[2]  L. Baxter‐Lowe,et al.  Two novel HLA-DQB1*06 alleles reveal additional heterogeneity of HLA-DQw1. , 1992, Tissue antigens.

[3]  J. Gorski,et al.  The predictive value of HLA-DR oligotyping for MLC responses. , 1992, Transplantation.

[4]  O. Olerup,et al.  HLA-DR typing by PCR amplification with sequence-specific primers (PCR-SSP) in 2 hours: an alternative to serological DR typing in clinical practice including donor-recipient matching in cadaveric transplantation. , 1992, Tissue antigens.

[5]  M. Carrington,et al.  Typing of HLA-DQA1 and DQB1 using DNA single-strand conformation polymorphism. , 1992, Human immunology.

[6]  L. Baxter‐Lowe,et al.  Characterization of a novel DQB1 allele associated with HLA-DQw3: implications for oligotyping. , 1992, Human immunology.

[7]  D. Stroncek.  Results of bone marrow transplants from unrelated donors , 1992, Transfusion.

[8]  M. Boyce-Jacino,et al.  HLA class II "typing": direct sequencing of DRB, DQB, and DQA genes. , 1992, Human immunology.

[9]  B. Mach,et al.  Selection of unrelated donors for bone marrow transplantation is improved by HLA class II genotyping with oligonucleotide hybridization. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[10]  R. T. D'Aquila,et al.  Maximizing sensitivity and specificity of PCR by pre-amplification heating , 1991, Nucleic Acids Res..

[11]  B. Mach,et al.  Genotypic typing of HLA class II: from the bench to the bedside. , 1991, Human immunology.

[12]  J. Bidwell,et al.  HLA‐DR/Dw MATCHING BY PCR FINGERPRINTING: THE ORIGIN OF PCR FINGERPRINTS AND FURTHER APPLICATIONS , 1991, European journal of immunogenetics : official journal of the British Society for Histocompatibility and Immunogenetics.

[13]  D. R. Lee,et al.  Solid-phase direct DNA sequencing of allele-specific polymerase chain reaction-amplified HLA-DR genes. , 1991, BioTechniques.

[14]  E. Thorsby,et al.  Distribution of HLA class II alleles among Norwegian Caucasians. , 1990, Human immunology.

[15]  B. Olaisen,et al.  Use of DNA amplification (PCR) and direct DNA sequencing in the characterization of C4 alleles , 1990, Annals of human genetics.

[16]  H. Inoko,et al.  A simple and rapid method for HLA-DRB and -DQB typing by digestion of PCR-amplified DNA with allele specific restriction endonucleases. , 1990, Tissue antigens.

[17]  H A Erlich,et al.  Genetic analysis of amplified DNA with immobilized sequence-specific oligonucleotide probes. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[18]  M. Uhlén,et al.  Direct solid phase sequencing of genomic and plasmid DNA using magnetic beads as solid support. , 1989, Nucleic acids research.

[19]  R. Higuchi,et al.  Simple and Rapid Preparation of Samples for PCR , 1989 .

[20]  H A Erlich,et al.  Generation of single-stranded DNA by the polymerase chain reaction and its application to direct sequencing of the HLA-DQA locus. , 1988, Proceedings of the National Academy of Sciences of the United States of America.

[21]  J. Devereux,et al.  A comprehensive set of sequence analysis programs for the VAX , 1984, Nucleic Acids Res..