Abstract
MLPA was developed by Schouten JP et al. in 2002, and has become a rapidly growing technique used in clinical diagnosis for genetic diseases, such as Duchenne muscular dystrophy (DMD)/Becker muscular dystrophy (BMD), Lynch syndrome and multiple congenital anomalies (MCA)/mental retardation (MR). MLPA is a simple and rapid method for simultaneous quantification of up to 40 nucleic acid sequences in a single reaction. MLPA probe consists of two different oligonucleotides, each containing one of the PCR primer sequences as well as a sequence complementary to the target sequence. The two probe oligonucleotides hybridize to immediately adjacent target sequences. It is only when the two probe parts are both hybridized to their target sequence that they can be ligated to each other by a thermostable ligase. These ligated probes will be amplified exponentially during PCR reaction. The resulting PCR amplification products are subsequently separated by capillary gel electrophoresis. The number of probe ligation products of one probe depends on the number of target sequences in the sample. The DMD gene is located on Xp21 and the gene responsible for DMD/BMD. Large deletions of DMD gene are found in 60%, large duplications are 10%, and small mutations are 30% of the cases. Although the conventional multiplex PCR assay was developed for the genetic testing of the DMD gene, it could not detect large duplication because of its low quantitative performance. The MLPA assay can provide a simple and rapid method for the large deletion and duplication screening done in DMD/BMD patients.
