The SALSA MLPA Probemix P190 CHEK2 is an in vitro diagnostic (IVD)1
or research use only (RUO) semi-quantitative assay2
for the detection of deletions or duplications in the human CHEK2
genes, and for the detection of the CHEK2
1100delC variant in genomic DNA isolated from human peripheral whole blood specimens. P190 CHEK2 is intended to confirm a potential cause for breast cancer in patients who are negative for BRCA1
mutations, and for other CHEK2
-related cancer types, including colorectal cancer. This product can also be used to determine increased cancer susceptibility in at-risk family members.
Copy number variations (CNVs) detected with P190 CHEK2 should be confirmed with a different technique. In particular, CNVs detected by only a single probe always require confirmation by another method. Most defects in the CHEK2
genes are point mutations, of which only the CHEK2
1100delC mutation will be detected by MLPA. It is therefore recommended to use this assay in combination with sequence analysis.
Please note that this probemix covers all exons of CHEK2
but not of ATM
. For the latter two genes, the P041/P042 ATM and P056 TP53 probemixes provide a better coverage and may detect aberrations that are not detected by this P190 CHEK2 probemix.
Assay results are intended to be used in conjunction with other clinical and diagnostic findings, consistent with professional standards of practice, including confirmation by alternative methods, clinical genetic evaluation, and counselling, as appropriate. The results of this test should be interpreted by a clinical molecular geneticist or equivalent.
This device is not intended to be used for standalone diagnostic purposes, pre-implantation or prenatal testing, population screening, or for the detection of, or screening for, acquired or somatic genetic aberrations, e.g. from DNA extracted from formalin-fixed paraffin embedded (FFPE) or fresh tumour materials.
Please note that this probemix is for in vitro diagnostic (IVD) use in the countries specified at the end of this product description. In all other countries, the product is for research use only (RUO).
To be used in combination with a SALSA MLPA Reagent Kit, Coffalyser.Net analysis software, and SALSA Binning DNA SD078.
all play important roles in DNA damage repair. A defect in one of these genes can lead to an increased risk of tumour formation. For breast cancer, autosomal dominant mutations in the genes BRCA1
are the most frequent cause, followed by mutations in CHEK2
, though with a much lower frequency (Buys et al. 2017). Mutations in CHEK2
may also increase the risk of developing colorectal cancer (Xiang et al. 2011) and other cancers, including prostate cancer (Cybulski et al. 2006). Mutations in CHEK2
, and the 1100delC mutation in particular, have also been suggested as an underlying cause of Li-Fraumeni syndrome (LFS) type 2. Moreover, a deletion in CHEK2
was found in a patient fulfilling Li-Fraumeni-Like (LFL) criteria (Ruijs et al. 2009). Researchers are uncertain whether CHEK2
mutations actually cause LFS or are merely associated with an increased risk of several types of cancer, including cancers seen in LFS. For more information: https://omim.org/entry/609265
exons 11-15 share a high sequence homology with several CHEK2
pseudogenes, which can result in a pseudogene-mediated gene conversion (Pan et al. 2017).
Autosomal dominant mutations in ATM
are linked to an increased risk of cancer, with breast cancer in particular (see Table 1). Autosomal recessive mutations of ATM
cause Ataxia-Telangiectasia, which is characterized by progressive cerebellar ataxia, telangiectases, and a predisposition to malignancy, particularly leukaemia and lymphoma. For more information: https://www.ncbi.nlm.nih.gov/books/NBK26468/
Autosomal dominant TP53
mutations result in LFS. The most common types of tumours in LFS are soft tissue sarcomas and osteosarcomas, pre-menopausal breast cancer, brain tumours, leukaemia, and adrenocortical carcinoma. Families presenting incomplete features of LFS are referred to as having LFL, and around 20-40% of these patients have a germline mutation in TP53
(Ruijs et al. 2010). More information on LFS: https://www.ncbi.nlm.nih.gov/books/NBK1311/
The SALSA MLPA Probemix P190-D1 CHEK2 contains 53 MLPA probes with amplification products between 124 and 500 nucleotides (nt). This includes 19 probes for the CHEK2
gene and one probe for the HSCB
gene, just upstream of CHEK2
. This probemix also contains one probe specific for the CHEK2
1100delC mutation, which will only generate a signal when the mutation is present. Moreover, 19 probes for ATM
and four probes for TP53
are also present. These probes target sequences in various parts of the genes, including in the first and last exon. In addition, ten 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 SD078
The SD078 Binning DNA provided with this probemix can be used for binning of all probes including one mutation-specific probe (313 nt probe 22034-SP0468-L31261 CHEK2
1100delC mutation). SD078 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 SD078 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. 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 SD078 Binning DNA product description, available online: www.mrcholland.com
Sample DNA developed for this product: