Multiplex Ligation-dependent Probe Amplification (MLPA) is a method to determine the copy number of up to 60 genomic DNA sequences in a single multiplex PCR based reaction. MLPA probes consist of two oligonucleotides that can hybridize next to each other on a certain DNA sequence of interest, where they are ligated. All ligated probes are subsequently amplified by PCR using a single set of primers. Each amplified MLPA probe has a unique length and can be visualized and quantified by capillary electrophoresis. As the primers are almost 100% consumed in the PCR reaction, the quantity of each PCR amplicon is proportional to the number of copies of each probe target sequence in the DNA sample. A trisomy 21 can therefore be detected by an approximately 50% increased signal of each chromosome 21 specific probe relative to reference samples. MLPA with the P095 Aneuploidy probemix for chromosomes 13, 18, 21, X and Y has been used as a rapid detection method on large numbers of samples from uncultured amniotic fluid or from chorionic villi. As compared to FISH and karyotyping, MLPA is more rapid, has a higher throughput, and is less expensive. MLPA however cannot detect low grade mosaicism, female triploidies, and copy number neutral chromosome abnormalities such as inversions and translocations.

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Methods in Molecular Biology
Department of Clinical Genetics

Schouten, J., van Vught, P., & Galjaard, R.-J. (2019). Multiplex ligation-dependent probe amplification (MLPA) for prenatal diagnosis of common aneuploidies. In Methods in Molecular Biology. doi:10.1007/978-1-4939-8889-1_11