An MLPA-based strategy for discrete CNV genotyping: CNV-miRNAs as an example

Copy number variation (CNV) has become well recognized in recent years. It has been estimated that common CNVs account for approximately 10% of the human genome and that they overlap hundreds of genes and other functional genetic elements. Although substantial progress in genome-wide CNV analysis has been made recently, there is still a need for a method that allows precise genotyping of selected CNVs. Here, we describe a novel strategy for CNV genotyping, taking advantage of the general principles of the multiplex ligation-dependent probe amplification (MLPA) method and short oligonucleotide probes, allowing easy custom design and generation of assays for almost any genomic region of interest. As a proof-of-concept, we developed two assays covering 17 candidate CNV regions that overlap human miRNA genes. Extensive quality control analysis demonstrated high reproducibility and reliability of the genotypes determined using our method. Detailed analysis of identified CNVs revealed that they are highly differentiated among the HapMap populations. The main advantages of the developed strategy include the simplicity of the assay design, its flexibility in terms of the selection of genomic regions, and its low cost (<$1-$10/genotype, depending on scale of experiment). These advantages make the presented strategy attractive for large-scale genetic analyses.