Complement C4 Gene Copy Number Variation Genotyping by High Resolution Melting PCR.

Claudia P Jaimes-Bernal,Francisco José Márquez,Monte Trujillo,Antonio Caruz

doi:10.3390/ijms21176309

Claudia P Jaimes-Bernal, Francisco José Márquez + Show 2 more

Open Access

https://doi.org/10.3390/ijms21176309

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Abstract

Background: Complement C4 gene copy number variation plays an important role as a determinant of genetic susceptibility to common diseases, such as systemic lupus erythematosus, schizophrenia, rheumatoid arthritis, and infectious diseases. This study aimed to develop an assay for the quantification of copy number variations in the C4 locus. Methods: the assay was based on a gene ratio analysis copy enumeration (GRACE) PCR combined with high resolution melting (HRM) PCR. The test was optimized using samples of a known genotype and validated with 72 DNA samples from healthy blood donors. Results: to validate the assay, standard curves were generated by plotting the C4/RP1 ratio values against copy number variation (CNV) for each gene, using genomic DNA with known C4 CNV. The range of copy numbers in control individuals was comparable to distributions observed in previous studies of European descent. Conclusions: the method herein described significantly simplifies C4 CNV diagnosis to validate the assay.

Highlights

The complement system is a key element of innate and acquired immunity, and acts as the first line of host defense
The C4 gene is located on the long arm of human chromosome 6 (6q21.3) in the major histocompatibility complex (MHC) class III region
The remaining 24% of the C4 genes do not have this retrovirus and are known as short C4 genes (C4S)

Summary

Introduction

The complement system is a key element of innate and acquired immunity, and acts as the first line of host defense. The C4 gene is located on the long arm of human chromosome 6 (6q21.3) in the major histocompatibility complex (MHC) class III region. This gene has two paralogs C4A and C4B, sharing 99% sequence identity, each of which is polymorphic in itself. These paralogs have different functional activities and affinities to antigenic surfaces. This study aimed to develop an assay for the quantification of copy number variations in the C4 locus. Results: to validate the assay, standard curves were generated by plotting the C4/RP1 ratio values against copy number variation (CNV) for each gene, using genomic DNA with known C4 CNV. Conclusions: the method described significantly simplifies C4 CNV diagnosis to validate the assay

Objectives

Methods

Results