A multiplex ligation‑dependent probe amplification‑based next‑generation sequencing approach for the detection of copy number variations in the human genome.

The aim of the present study was to describe a multiplex ligation-dependent probe amplification (MLPA)-based next-generation sequencing (NGS) assay that exhibited a significantly higher efficiency in detecting copy number variations (CNVs) and known single-nucleotide variants, compared with traditional MLPA. MLPA polymerase chain reaction products were used to construct a library with indexed adapters, which was subsequently tested on an NGS platform, and the resulting data were analyzed by a series of analytical software. The reads from each probe reflected genetic variations in the target regions, and fragment differentiation was based on the specific base composition of the sequences, rather than fragment length, which was determined by capillary electrophoresis. The results of this approach were not only consistent with the MLPA results following capillary electrophoresis, but also coincided with the CNV results from the single-nucleotide polymorphism array chip. This method allowed high-throughput screening for the number of fragments and samples by integrating additional indices for detection. Furthermore, this technology precisely and accurately performed large-scale detection and quantification of DNA variations, thereby serving as an effective and sensitive method for diagnosing genetic disorders caused by CNVs and known single-nucleotide variations. Notably, MLPA-NGS circumvents the problems associated with the inaccuracies of NGS in CNV detection due to the use of target sequence capture.