Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
2
pubmed:dateCreated
1997-2-4
pubmed:abstractText
Molecular typing of HLA DQB1 alleles, employing sequence-specific primers (SSP) for PCR amplification, was used to test a novel method that eliminates the requirement for subsequent gel electrophoresis or additional hybridization steps by directly detecting positive reactions. We have evaluated the performance of this fluorescence-based oligonucleotide probe assay to assign the most common DQB1 alleles on DNA from 14 homozygous cell lines and in a blind study of 50 diabetic patient samples that had been previously typed at the DQB1 locus using SSOP and conventional SSP-based approaches. We used a panel of 14 DQB1 SSP primer pairs, internal control primers, and a combination of 4 fluorescent oligonucleotide probes to detect 14 alleles or groups of alleles and controls. We can reliably detect single-base allelic differences, observe 100% concordance with the results obtained using both of the standard methods, and are able to further subtype several alleles that are not easily distinguished using SSOP (e.g. DQB1 *0401/0402 and DQB1 *0302/ 0303). Sequence-specific priming and exonuclease-released fluorescence (SSPERF) detection is technically simple and can be performed in less than 2 hours, including DNA extraction, PCR amplification, data analysis and allele identification. This method is particularly useful for the analysis of large numbers of samples, for which high throughput is critical and for which gel-based approaches are difficult to perform. This technique may also be useful for small-scale class I and class II molecular typing in clinically oriented laboratories.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Aug
pubmed:issn
0001-2815
pubmed:author
pubmed:issnType
Print
pubmed:volume
48
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
97-112
pubmed:dateRevised
2011-11-17
pubmed:meshHeading
pubmed:year
1996
pubmed:articleTitle
Sequence-specific priming and exonuclease-released fluorescence detection of HLA-DQB1 alleles.
pubmed:affiliation
Department of Pediatrics, University of Pittsburgh, School of Medicine, Pennsylvania, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, Non-P.H.S.