In:
Tissue Antigens, Wiley, Vol. 44, No. 3 ( 1994-09), p. 137-147
Abstract:
Abstract: We have developed a simple and rapid method for DNA typing of the HLA‐A locus using PCR amplification and hybridization of the PCR product, labeled with biotinylated primers, to an array of immobilized oligonucleotide probes in a single hybridization reaction (reverse dot or line blot). A single primer set (RAP1007 and DB337) is used to specifically amplify a 990‐bp fragment containing the HLA‐A locus exons 1, 2, and 3 from genomic DNA. This primer set is locus‐specific and amplifies all HLA‐A alleles. A set of 51 sequence‐specific oligonucleotide (SSO) probes, 25 for exon 2 and 26 for exon 3, was immobilized to a nylon membrane by UV‐crosslinking oligonucleotide probes containing a poly‐thymidine “tail” added with terminal transferase. In the line blot format, all 50 SSO probes plus a control probe are immobilized on a single nylon membrane strip. The probe array was used for typing in a hybridization reaction with DNA amplified from a variety of samples. These probes can identify 37 homozygous HLA‐A alleles. In the analysis of heterozygous samples, 604 heterozygous types out of 633 (95.4%) possible heterozygous probe patterns can be detected as a unique probe reactivity pattern. A simple computer program has been developed to assign the alleles and genotypes based on the probe hybridization pattern.
Type of Medium:
Online Resource
ISSN:
0001-2815
,
1399-0039
DOI:
10.1111/tan.1994.44.issue-3
DOI:
10.1111/j.1399-0039.1994.tb02371.x
Language:
English
Publisher:
Wiley
Publication Date:
1994
detail.hit.zdb_id:
1498140-3
detail.hit.zdb_id:
2844321-4
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