title: Genotyping for Glycophorin GYP(B-A-B) Hybrid Genes Using a Single Nucleotide Polymorphism-Based Algorithm by Matrix-Assisted Laser Desorption/Ionisation, Time-of-Flight Mass Spectrometry.
abstract: The genetic basis for five GP(B-A-B) MNS system hybrid glycophorin blood group antigens results from rearrangement between the homologous GYPA and GYPB genes. Each hybrid glycophorin displays a characteristic profile of antigens. Currently, no commercial serological reagents are currently available to serologically type for these antigens. The aim of this study was to develop a single nucleotide polymorphism (SNP) mapping genotyping technique to allow characterisation of various GYP(B-A-B) hybrid alleles. Matrix-assisted laser desorption/ionisation time-of-flight (MALDI-TOF) mass spectrometry (MS) assays were designed to genotype five GYP(B-A-B) hybrid alleles. Eight nucleotide positions were targeted and incorporated into the SNP mapping protocol. The allelic frequencies were calculated using peak areas. Sanger sequencing was performed to resolve a GYP*Hop 3' breakpoint. Observed allelic peak area ratios either coincided with the expected ratio or were skewed (above or below) from the expected ratio with switching occurring at and after the expected break point to generate characteristic mass spectral plots for each hybrid. Sequencing showed that the GYP*Hop crossover in the intron 3 region, for this example, was identical to that for GYP*Bun reference sequence. An analytical algorithm using MALDI-TOF MS genotyping platform defined GYPA inserts for five GYP(B-A-B) hybrids. The SNP mapping technique described here demonstrates proof of concept that this technology is viable for genotyping hybrid glycophorins, GYP(A-B-A), GYP(A-B) and GYP(B-A), and addresses the gap in current typing technologies.
authors.affiliation: Clinical Services and Research, Australian Red Cross Blood Service, 44 Musk Avenue, Kelvin Grove, QLD, 4059, Australia.|Institute of Clinical Blood Transfusion, Guangzhou Blood Center, Guangzhou, People's Republic of China.
authors.forename: Ling
authors.initials: L
authors.lastname: Wei
authors.affiliation: Clinical Services and Research, Australian Red Cross Blood Service, 44 Musk Avenue, Kelvin Grove, QLD, 4059, Australia.
authors.forename: Genghis H
authors.initials: GH
authors.lastname: Lopez
authors.affiliation: Institute of Clinical Blood Transfusion, Guangzhou Blood Center, Guangzhou, People's Republic of China.
authors.forename: Yanli
authors.initials: Y
authors.lastname: Ji
authors.affiliation: Red Cell Reference Laboratory, Australian Red Cross Blood Service, 100-154 Batman St., West Melbourne, VIC, 3003, Australia.
authors.forename: Jennifer A
authors.initials: JA
authors.lastname: Condon
authors.affiliation: Agena Bioscience, Herston, QLD, 4006, Australia.
authors.forename: Darryl L
authors.initials: DL
authors.lastname: Irwin
authors.affiliation: Institute of Clinical Blood Transfusion, Guangzhou Blood Center, Guangzhou, People's Republic of China.
authors.forename: Guangping
authors.initials: G
authors.lastname: Luo
authors.affiliation: Clinical Services and Research, Australian Red Cross Blood Service, 44 Musk Avenue, Kelvin Grove, QLD, 4059, Australia.
authors.forename: Catherine A
authors.initials: CA
authors.lastname: Hyland
authors.affiliation: Clinical Services and Research, Australian Red Cross Blood Service, 44 Musk Avenue, Kelvin Grove, QLD, 4059, Australia. rFlower@redcrossblood.org.au.
authors.forename: Robert L
authors.initials: RL
authors.lastname: Flower
mesh_terms: D000465:Algorithms; D005787:Gene Frequency; D060005:Genotyping Techniques / Q000379:methods*; D006021:Glycophorins / Q000235:genetics*; D006801:Humans; D020641:Polymorphism, Single Nucleotide; D017422:Sequence Analysis, DNA; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods
chemical_list: C409972:GYPA protein, human; C409973:GYPB protein, human; D006021:Glycophorins
keywords: GP(B-A-B) hybrid glycophorin; GYP(B-A-B) genotyping; MALDI-TOF MS genotyping