Copy Number Variation Analysis Based on Gold Magnetic Nanoparticles and Fluorescence Multiplex Ligation-Dependent Probe Amplification

A novel strategy for determining relative copy number of target genes was developed in this study based on gold magnetic nanoparticles (GMNPs) and fluorescence multiplex ligation-dependent probe amplification (fMLPA) assay. This method relies on the following stages: (1) amplification of target and reference genes by fMLPA, (2) magnetic capture of fMLPA products on GMNPs, (3) detection of the nanocomplexes by fluorescence scanner, (4) relative quantification of target gene, (5) calculation of relative copy number. The trisomy 21-related Decoy Receptor 3 (DSCR3) gene and reference glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene were selected for testing this technique. Under optimized parameters, results from 40 clinical samples showed equivalent relative copy numbers compared to theoretical values. This promising approach is simple, sensitive, selective and with low fluorescent background, thus making it suitable for copy number variation (CNV) analysis, with potential for diagnosis of genetic diseases.