General information
The SALSA MLPA
Probemix P480 WHS & Achondroplasia is a
research use only (RUO) assay for the detection of deletions or duplications in the 4p16 chromosomal region, which is associated with Wolf-Hirschhorn syndrome (WHS). This probemix can also be used to detect the presence of the c.1138G>A (p.Gly380Arg) point mutation, which causes the vast majority of cases of achondroplasia (ACH).
WHS is characterised by severe growth retardation, intellectual disability of variable degrees, microcephaly, “Greek helmet” facies, and closure defects (cleft lip or palate, coloboma of the eye, and cardiac septal defects), among others (Zollino et al. 2008). The prevalence of this syndrome is estimated at about 1 in 50,000 births, although it is suspected to be higher, at about 1 in 20,000 births (Berrocoso et al. 2020). Furthermore, the incidence is twice as high in females than males (Coles et al. 1992). The syndrome is caused by a heterozygous deletion of the WHS critical region on 4p16.3. Most individuals with WHS have
de novo 4p terminal and interstitial deletions (50-60%),
de novo microdeletions (25-30%), or an unbalanced translocation (~15%). The remainder have other complex rearrangements leading to 4p16 deletion (South et al. 2008). The deletion extent of the 4p subtelomeric region varies between patients, with smaller deletions resulting in a milder phenotype than larger ones (Zollino et al. 2008).
One of the genes in the 4p16 chromosomal region is the
FGFR3 gene. The
c.1138G>A mutation in exon 10 of the
FGFR3 gene causes approximately 98% of cases of ACH (Etlik et al. 2008). ACH is the most common cause of disproportionate short stature. The worldwide prevalence of this genetic disorder is estimated at about 1 in 22,000 births (Foreman et al. 2020).
More information is available at
https://www.ncbi.nlm.nih.gov/books/NBK1183/ (WHS) and
https://www.ncbi.nlm.nih.gov/books/NBK1152/ (ACH).
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 P480-A1 WHS & Achondroplasia contains 51 MLPA probes with amplification products between 121 and 505 nucleotides (nt). This includes 38 probes for the 4p16
chromosomal region. Furthermore, this probemix also contains one probe specific for the
c.1138G>A mutation in
FGFR3 which will only generate a signal when the mutation is present. In addition, 12 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.mrcholland.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, and 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.mrcholland.com.
SALSA Binning DNA SD080
The SD080 Binning DNA provided with this probemix can be used for binning of all probes including the mutation-specific probe (probe 21987-L30825 for the
FGFR3 c.1138G>A mutation). SD080 Binning DNA is a mixture of genomic DNA from healthy individuals and synthetic DNA that contains the target sequence detected by the above mentioned probe. Inclusion of one reaction with 5 μl SD080 Binning DNA in initial MLPA experiments is essential as it can be used to aid in data binning of the peak pattern using Coffalyser.Net software. Furthermore, Binning DNA should be included in the experiment whenever changes have been applied to the set-up of the capillary electrophoresis device (e.g. when capillaries have been renewed). Binning DNA should never be used as a reference sample in the MLPA data analysis, neither should it be used in quantification of mutation signal(s). It is strongly advised that all samples tested are extracted with the same method and derived from the same source of tissue. For further details, please consult the SD080 Binning DNA product description, available online:
www.mrcholland.com.
This product is for research use only (RUO).
Sample DNA
Sample DNA developed for this product: