Concordance of next generation sequence-based and sequence specific oligonucleotide probe-based HLA-DRB1 genotyping

Julie A. Lane, Jameel R. Johnson, Janelle A. Noble

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

Next generation sequencing (NGS) of clonally amplified DNA, using Roche 454 technology, was used to genotype HLA-DRB1, DRB3, DRB4, and DRB5 loci (exon 2 only) from a set of 993 samples from newborns with maternally-reported African American ancestry. DRB1 exon 2 was genotyped previously on the same sample set using sequence-specific oligonucleotide probe (SSOP) technology. Comparison of the genotype calls from both methods indicated concordance of 92.3%. Some discordance was expected due to the higher resolution of NGS data, compared to SSOP data. This resulted from selection of the incorrect allele from the ambiguity string produced by SSOP genotyping. Of 76 discordant genotypes, only three were due to resolution of ambiguity with the NGS method. The low percent of changes due to the increased resolution of the NGS method instills confidence in the overall value of previous data genotyped with moderate resolution methods, i.e., the vast majority of alleles present in a population are those that are detectable at moderate resolution. The remaining 73 discordant genotypes resulted from preventable errors in sample handling, data interpretation, and data entry. These results underscore the potential for error that can result from factors such as low quality genomic DNA, manual data entry, and interpretation of marginal genotyping results. Optimization of genomic DNA quality, automation of genotyping steps wherever possible, and use of the highest resolution technology available can lead to dramatic improvements in HLA genotype data quality. NGS-based methodology generated data of superior quality and accuracy compared to the SSOP system.

Original languageEnglish (US)
Pages (from-to)939-944
Number of pages6
JournalHuman Immunology
Volume76
Issue number12
DOIs
StatePublished - Dec 1 2015
Externally publishedYes

Fingerprint

HLA-DRB1 Chains
Oligonucleotide Probes
Genotype
Technology
HLA-DRB3 Chains
Exons
DNA
Alleles
Automation
African Americans
Newborn Infant
Population

Keywords

  • DRB genotyping
  • HLA
  • Next-generation sequencing

ASJC Scopus subject areas

  • Immunology
  • Immunology and Allergy

Cite this

Concordance of next generation sequence-based and sequence specific oligonucleotide probe-based HLA-DRB1 genotyping. / Lane, Julie A.; Johnson, Jameel R.; Noble, Janelle A.

In: Human Immunology, Vol. 76, No. 12, 01.12.2015, p. 939-944.

Research output: Contribution to journalArticle

Lane, Julie A. ; Johnson, Jameel R. ; Noble, Janelle A. / Concordance of next generation sequence-based and sequence specific oligonucleotide probe-based HLA-DRB1 genotyping. In: Human Immunology. 2015 ; Vol. 76, No. 12. pp. 939-944.
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