General information
SALSA
® MLPA
® Probemix P105 Glioma is a
research use only (RUO) assay for the detection of deletions or gains in the following genes
PDGFRA (4q12),
EGFR (7p11.2),
CDKN2A (9p21.3),
PTEN (10q23.31),
CDK4-MIR26A2-MDM2 (12q14-q15),
NFKBIA (14q13.2) and
TP53 (17p13.1). Moreover, this probemix can be used to detect the chr. 7 gains and chr. 10 losses and to detect the presence of
TERT promoter mutations C228T and C250T.
Gliomas are the most common primary brain tumours and account for one third of central nervous system (CNS) tumours. Gliomas comprise a very heterogeneous group of CNS neoplasms derived from glial cells. There are several oncogenes and tumour suppressor genes, which have been shown to undergo copy number changes in gliomas. Somatic mutations, disruptions, or copy number aberrations in three critical signalling pathways, a) the RTK/PI3K pathway (involving e.g.
EGFR, PDGFRA and
PTEN genes), b) the p53 pathway (involving e.g.
CDKN2A, MDM2 and
TP53 genes) and c) the RB pathway (involving e.g.
CDKN2A and
CDK4 genes), are suggested to contribute to the development of gliomas (Cancer Genome Atlas Research Network 2008).
Epidermal growth factor receptor (EGFR) and its ligands are cell signalling molecules involved in diverse cellular functions. These include cell proliferation, differentiation, motility and survival, and tissue development. Glioblastomas often express EGFR variant III (EGFRvIII), a constitutively active genomic deletion variant of
EGFR which is characterised by deletions of exons 2-7 of the
EGFR gene (Sugawa et al. 1990). This probemix allows detection of deletions of
EGFR that result in EGFRvIII.
Point mutations in
TERT promoter region generate novel transcription factor binding sites and thus increase the expression of telomerase enzyme encoded by
TERT. Common
TERT promoter mutations are known as C228T and C250T, referring to C>T transitions at hg19/GCRh37 chr5:1295
228 and chr5:1295
250 positions, respectively. These mutations are predominantly present in oligodendroglioma and are associated with poor prognosis and reduced survival in the absence of
IDH-mutation (Labussière et al. 2014).
TERT promoter mutation, in combination with
IDH-mutation and 1p/19q codeletion, is characteristic of oligodendroglioma. Absence of
TERT promoter mutation, coupled with the presence of
IDH-mutation, designates astrocytoma (Cahill et al. 2015; Eckel-Passow et al. 2015).
This product is not CE/FDA registered for use in diagnostic procedures. The SALSA® MLPA® technique is covered by US patent 6,955,901 and corresponding patents outside the US. The purchase of this product includes a license to use only this amount of product solely for the purchaser’s own use.
Probemix content
The SALSA MLPA Probemix P105-E1 Glioma contains 60 MLPA probes with amplification products between 120 and 500 nucleotides (nt). This includes in total 43 probes for the
PDGFRA,
EGFR,
CDKN2A,
PTEN,
CDK4, MIR26A2, MDM2,
NFKBIA and
TP53 genes, and single flanking probes for 7q and 10p arms
. Furthermore, this probemix contains two probes specific for the
TERT C228T and C250T mutations, which will only generate a signal when the mutation is present. In addition, 13 reference probes are included that detect relatively copy number stable regions in various cancer types including gliomas. Complete probe sequences and the identity of the genes detected by the reference probes are available in the product description and 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 SD097
SALSA
® Binning DNA SD097 provided with this probemix can be used for binning of all probes including the two mutation-specific probes (126 nt probe S1310-L32991 for
TERT C250T and 156 nt probe 23341-L33133 for
TERT C228T). SD097 is a mixture of human female genomic DNA from healthy individuals and a titrated amount of plasmid DNA that contains the target sequence detected by the above mentioned probes. Inclusion of one reaction with 5 μl SD097 in initial MLPA experiments is essential as it can be used to aid in data binning of the peak pattern using Coffalyser.Net. 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 signals. For further details, please consult the SD097 product description, available online:
www.mrcholland.com.
This product is for research use only (RUO).
Sample DNA
Sample DNA developed for this product: