General information
The SALSA MLPA Probemix P222 LCA mix-2 is a research use only (RUO) assay for the detection of deletions or duplications in the GUCY2D, RDH12 and RPGRIP1 genes, which are associated with Leber Congenital Amaurosis (LCA). This probemix can also be used to detect the presence of the c.2991+1655A>G (rs281865192; p.Cys998X) point mutation in CEP290, which is also linked to LCA.

LCA comprises of a group of early-onset childhood retinal dystrophies and is characterised by vision loss, nystagmus and severe retinal dysfunction. It is the most common inherited cause of blindness. Pathogenic variants in at least seventeen genes are known to cause LCA. This probemix contains probes for four of these genes.

The protein encoded by the GUCY2D gene is retina-specific guanylate cyclase, located in the photoreceptor cells where it is involved in phototransduction. Defects in the GUCY2D gene account for 6-21% of all LCA cases. The protein encoded by the RDH12 gene is an NADPH-dependent retinal reductase, which metabolizes both trans- and cis-retinols. Defects in the RDH12 gene account for 4-10% of all LCA cases. The protein encoded by the RPGRIP1 gene is retinitis pigmentosa GTPase regulator interacting protein, a key component of photoreceptor cells. Defects in the RPGRIP1 gene account for ~5% of all LCA cases. The protein encoded by the CEP290 gene is centrosomal protein of 290 kDa, involved in ciliary assembly and ciliary trafficking of the photoreceptor cells. Defects in the CEP290 gene account for 15%-22% of all LCA cases. The intronic mutation c.2991+1655A>G is the most common CEP290 mutation associated with LCA and may account for up to 21% of LCA cases (den Hollander et al. 2006).

More information is available at https://www.ncbi.nlm.nih.gov/books/NBK531510/.

This SALSA MLPA probemix is not CE/FDA registered for use in diagnostic procedures. Purchase of this product includes a limited license for research purposes.

Probemix content
The SALSA MLPA Probemix P222-B2 LCA mix-2 contains 44 MLPA probes with amplification products between 130 and 454 nucleotides (nt). This includes 18 probes for the RPGRIP1 gene, one probe for each exon with the exception of exons 5, 7, 9, 12, 15 and 17, seven probes for the RDH12 gene, one probe for each exon with the exception of exons 1 and 2, and eight probes for the GUCY2D gene, one probe each for exons 2, 5, 7, 10, 12, 14, 17 and 20. Furthermore, this probemix also contains one probe specific for the c.2991+1655A>G mutation in the CEP290 gene, which will only generate a signal when the wildtype allele is present. A reduced signal can point towards the presence of the mutation or a deletion in intron 26. To facilitate discrimination between presence of the mutation or presence of a deletion, one additional probe for intron 26 of CEP290 is included in the probemix. This probe is located at short distance from the wild-type probe. In case of a deletion, both probes are expected so show a reduced signal. In case the mutation is present, only the probe specific for the the c.2991+1655A>G mutation is expected to show a reduced signal. In addition to the above listed probes, nine reference probes are included that detect autosomal chromosomal locations. Complete probe sequences and the identity of the genes detected by the reference probes are available online (www.mlpa.com).

This probemix contains nine quality control fragments generating amplification products between 64 and 105 nt: four DNA Quantity fragments (Q-fragments), two DNA Denaturation fragments (D-fragments), one Benchmark fragment, one chromosome X and one chromosome Y-specific fragment. More information on how to interpret observations on these control fragments can be found in the MLPA General Protocol and online at www.mlpa.com.