HLA-DQB1 genotyping with simple automated DNA sequencing and single-strandconformation polymorphism analysis

Citation
Ch. Yang et al., HLA-DQB1 genotyping with simple automated DNA sequencing and single-strandconformation polymorphism analysis, J FORMOS ME, 99(9), 2000, pp. 698-703
Citations number
16
Categorie Soggetti
General & Internal Medicine
Journal title
JOURNAL OF THE FORMOSAN MEDICAL ASSOCIATION
ISSN journal
09296646 → ACNP
Volume
99
Issue
9
Year of publication
2000
Pages
698 - 703
Database
ISI
SICI code
0929-6646(200009)99:9<698:HGWSAD>2.0.ZU;2-T
Abstract
Background and purpose: Accurate human leukocyte antigen (HLA) typing is im portant for matching donors and recipients of bone marrow transplantation. Because HLA is highly polymorphic, HLA genotyping is also a valuable tool i n forensic identification of humans. The primary objective of this study wa s to establish a simple, rapid, and economic HLA analysis system suitable f or use in forensic applications. Method: We used the primer pair DB130 and GH29 to amplify the HLA class II DQB1 gene by polymerase chain reaction (PCR). The nucleotide sequences of t he PCR products were analyzed with an ABI Prism 377 automatic DNA sequencer , with the aid of its systematic analytical procedure. Some genotypes were confirmed by single-strand conformation polymorphism (SSCP) analysis. Results: We identified 15 alleles and 37 genotypes from 86 Taiwanese subjec ts. The most frequent allele was 03011 (27.9%) and the most frequent genoty pe was 03011/03011 (15.1%). Statistical analysis showed that the allelic di versity was 0.862 and the power of discrimination was 0.948. Conclusions: The results of this study indicate that the combined use of au tomated sequencing with only one primer pair and SSCP provides a simple, ra pid, and economic tool for analyzing the DQB1 gene. Compared with other seq uencing methods that use a set of multiple primers, this method has two adv antages. First, it is simpler and faster, because the HLAB1 genotype can be determined in a single PCR reaction. Second, the use of only one primer se t obviates the need for preferential annealing of any one primer set.