title: A comparison of assays for accurate copy number measurement of the low-affinity Fc gamma receptor genes FCGR3A and FCGR3B.
abstract: The FCGR3 locus encoding the low affinity activating receptor FcγRIII, plays a vital role in immunity triggered by cellular effector and regulatory functions. Copy number of the genes FCGR3A and FCGR3B has previously been reported to affect susceptibility to several autoimmune diseases and chronic inflammatory conditions. However, such genetic association studies often yield inconsistent results; hence require assays that are robust with low error rate. We investigated the accuracy and efficiency in estimating FCGR3 CNV by comparing Sequenom MassARRAY and paralogue ratio test-restriction enzyme digest variant ratio (PRT-REDVR). In addition, since many genetic association studies of FCGR3B CNV were carried out using real-time quantitative PCR, we have also included the evaluation of that method's performance in estimating the multi-allelic CNV of FCGR3B. The qPCR assay exhibited a considerably broader distribution of signal intensity, potentially introducing error in estimation of copy number and higher false positive rates. Both Sequenom and PRT-REDVR showed lesser systematic bias, but Sequenom skewed towards copy number normal (CN = 2). The discrepancy between Sequenom and PRT-REDVR might be attributed either to batch effects noise in individual measurements. Our study suggests that PRT-REDVR is more robust and accurate in genotyping the CNV of FCGR3, but highlights the needs of multiple independent assays for extensive validation when performing a genetic association study with multi-allelic CNVs.
authors.affiliation: Institute of Medical Molecular Biotechnology (IMMB), Faculty of Medicine, Universiti Teknologi MARA, Sungai Buloh Campus, Selangor, Malaysia.
authors.forename: Umi Shakina
authors.initials: US
authors.lastname: Haridan
authors.affiliation: Institute of Medical Molecular Biotechnology (IMMB), Faculty of Medicine, Universiti Teknologi MARA, Sungai Buloh Campus, Selangor, Malaysia.
authors.forename: Umairah
authors.initials: U
authors.lastname: Mokhtar
authors.affiliation: Department of Genetics, University of Leicester, Leicester, United Kingdom.
authors.forename: Lee R
authors.initials: LR
authors.lastname: Machado
authors.affiliation: Institute of Medical Molecular Biotechnology (IMMB), Faculty of Medicine, Universiti Teknologi MARA, Sungai Buloh Campus, Selangor, Malaysia.
authors.forename: Abu Thalhah
authors.initials: AT
authors.lastname: Abdul Aziz
authors.affiliation: Department of Microbiology and Parasitology, School of Medical Science, Health Campus, Universiti Sains Malaysia, Kubang Kerian, Kelantan, Malaysia.
authors.forename: Rafidah Hanim
authors.initials: RH
authors.lastname: Shueb
authors.affiliation: Department of Medicine, Hospital Sungai Buloh, Jalan Hospital, Sungai Buloh, Selangor, Malaysia.
authors.forename: Masliza
authors.initials: M
authors.lastname: Zaid
authors.affiliation: Department of Medicine, Hospital Sungai Buloh, Jalan Hospital, Sungai Buloh, Selangor, Malaysia.
authors.forename: Benedict
authors.initials: B
authors.lastname: Sim
authors.affiliation: Hospital Raja Perempuan Zainab II, Kota Bharu, Kelantan, Malaysia.
authors.forename: Mahiran
authors.initials: M
authors.lastname: Mustafa
authors.affiliation: Hospital Raja Perempuan Zainab II, Kota Bharu, Kelantan, Malaysia.
authors.forename: Nik Khairudin
authors.initials: NK
authors.lastname: Nik Yusof
authors.affiliation: Department of Medicine, Hospital Sungai Buloh, Jalan Hospital, Sungai Buloh, Selangor, Malaysia.
authors.forename: Christopher K C
authors.initials: CK
authors.lastname: Lee
authors.affiliation: Department of Biomedical Science, Faculty of Medicine and Health Science, Universiti Putra Malaysia, Serdang, Selangor, Malaysia.
authors.forename: Suhaili
authors.initials: S
authors.lastname: Abu Bakar
authors.affiliation: Department of Medical Microbiology, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia; Tropical Infectious Disease Research and Education Centre, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia.
authors.forename: Sazaly
authors.initials: S
authors.lastname: AbuBakar
authors.affiliation: Department of Genetics, University of Leicester, Leicester, United Kingdom.
authors.forename: Edward J
authors.initials: EJ
authors.lastname: Hollox
authors.affiliation: Institute of Medical Molecular Biotechnology (IMMB), Faculty of Medicine, Universiti Teknologi MARA, Sungai Buloh Campus, Selangor, Malaysia.
authors.forename: Hoh
authors.initials: H
authors.lastname: Boon Peng
mesh_terms: D001681:Biological Assay / Q000379:methods*; D016022:Case-Control Studies; D056915:DNA Copy Number Variations / Q000235:genetics; D003715:Dengue / Q000235:genetics; D058851:GPI-Linked Proteins / Q000235:genetics; D020022:Genetic Predisposition to Disease / Q000235:genetics; D005838:Genotype; D006801:Humans; D060888:Real-Time Polymerase Chain Reaction; D017452:Receptors, IgG / Q000235:genetics*
chemical_list: C496124:FCGR3A protein, human; C428200:FCGR3B protein, human; D058851:GPI-Linked Proteins; D017452:Receptors, IgG