SALSA MLPA P090 BRCA2 probemix
Intended use: The SALSA MLPA probemix P090 BRCA2 is an in vitro diagnostic (IVD)1 or a research use only (RUO) assay for the detection of deletions or duplications in the human BRCA2 gene, and the presence of the c.156_157insAlu mutation, in order to confirm a potential cause and clinical diagnosis for hereditary breast and ovarian cancer (HBOC). This product can also be used for molecular genetic testing of at-risk family members.
This assay is for use with human DNA extracted from peripheral blood and not for use with DNA extracted from formalin-fixed paraffin embedded or fresh tumour materials. Deletions or duplications detected with the P090 BRCA2 probemix must be verified by using the SALSA MLPA probemix P077 BRCA2 Confirmation or a different technique. Most defects in the BRCA2 gene are point mutations, the majority of which will not be detected by MLPA. It is therefore recommended to use this SALSA MLPA probemix in combination with sequence analysis of the BRCA2 gene. This assay is not intended to be used as a standalone assay for clinical decisions. The results of this test should be interpreted by a clinical molecular geneticist or equivalent.
1Please note that this probemix is for In Vitro Diagnostic use (IVD) in the countries specified at the end of this product description. In all other countries, the product is for Research Use Only (RUO).
Clinical background: Breast and ovarian carcinomas are among the most common malignancies in developed countries. The majority of cases are considered sporadic, but in a substantial portion, a clear history of cases within a family is present. The BRCA1 and BRCA2 proteins are associated with the activation of double-strand break repair and homologous recombination and are important in maintaining genomic stability. Germline mutations in the BRCA1 and BRCA2 genes are linked to a high risk of young-onset hereditary breast and ovarian cancer. Features characteristic of hereditary, versus sporadic, breast cancer are: younger age at diagnosis, frequent bilateral disease, and more frequent occurrence of disease among male relatives. Mutations in the BRCA1 and BRCA2 genes account for about 20 to 25% of hereditary breast cancers (Easton 1999) and about 5 to 10% of all breast cancers (Campeau et al. 2008). In addition, mutations in the BRCA1 and BRCA2 genes cause around 15% of ovarian cancers overall (Pal et al. 2005). More information is available at http://www.ncbi.nlm.nih.gov/books/NBK1247/.
Deletions or duplications are more frequent for BRCA1 than for BRCA2. The prevalence of deletions or duplications is dependent on the studied population and ranges from 0% to 11% of all BRCA2 mutations (Agata et al. 2005, Woodward et al. 2005, Casilli et al. 2006, Stadler et al. 2010).
P090-B1 probemix content: This SALSA MLPA probemix P090-B1 BRCA2 contains 51 MLPA probes with amplification products between 130 and 499 nt including 40 probes for the BRCA2 gene region and 11 reference probes that detect sequences outside this region. The identity of the genes detected by the reference probes is available online (www.mlpa.com). At least one MLPA probe is present for each exon in the BRCA2 transcript; two probes are present for exons 1 and 3, three probes are present for exons 10 and 27, and six probes are present for exon 11. One of the probes for exon 3 detects the wild type sequence of the c.156_157insAlu mutation and a reduced signal can point towards the presence of this mutation or a deletion of exon 3. In addition, there is a probe for the sequence upstream and a probe for the sequence downstream of BRCA2.
This probemix contains nine quality control fragments generating amplification products between 64 and 105 nt: four DNA Quantity Fragments (Q-fragments), three DNA Denaturation Fragments (D-fragments), and one chromosome X and one chromosome Y-specific fragment. The Q-fragments are only visible when less than 100 ng sample DNA is used. Low signal of the 88 and 96 nt fragment indicates incomplete DNA denaturation. More information on how to interpret observations on these control fragments can be found in the MLPA General Protocol.
Positive control DNA samples: In case no positive DNA sample is available in your laboratory, an artificial duplication DNA sample for this probemix (catalogue number SD024) can be ordered from MRC-Holland. This SD024 Artificial Duplication DNA will show a duplication of several probes when using the following probemixes: P002 and P087 BRCA1; P045, P090 and P077 BRCA2. The SD024 Artificial Duplication DNA is a mixture of human female genomic DNA and a titrated amount of plasmid containing selected probe target sequences. For further details, please consult the SD024 Artificial Duplication DNA product description, available online: www.mlpa.com. This SD024 is for research use only (RUO).
Sample DNA
Sample DNA developed for this product:
This assay is for use with human DNA extracted from peripheral blood and not for use with DNA extracted from formalin-fixed paraffin embedded or fresh tumour materials. Deletions or duplications detected with the P090 BRCA2 probemix must be verified by using the SALSA MLPA probemix P077 BRCA2 Confirmation or a different technique. Most defects in the BRCA2 gene are point mutations, the majority of which will not be detected by MLPA. It is therefore recommended to use this SALSA MLPA probemix in combination with sequence analysis of the BRCA2 gene. This assay is not intended to be used as a standalone assay for clinical decisions. The results of this test should be interpreted by a clinical molecular geneticist or equivalent.
1Please note that this probemix is for In Vitro Diagnostic use (IVD) in the countries specified at the end of this product description. In all other countries, the product is for Research Use Only (RUO).
Clinical background: Breast and ovarian carcinomas are among the most common malignancies in developed countries. The majority of cases are considered sporadic, but in a substantial portion, a clear history of cases within a family is present. The BRCA1 and BRCA2 proteins are associated with the activation of double-strand break repair and homologous recombination and are important in maintaining genomic stability. Germline mutations in the BRCA1 and BRCA2 genes are linked to a high risk of young-onset hereditary breast and ovarian cancer. Features characteristic of hereditary, versus sporadic, breast cancer are: younger age at diagnosis, frequent bilateral disease, and more frequent occurrence of disease among male relatives. Mutations in the BRCA1 and BRCA2 genes account for about 20 to 25% of hereditary breast cancers (Easton 1999) and about 5 to 10% of all breast cancers (Campeau et al. 2008). In addition, mutations in the BRCA1 and BRCA2 genes cause around 15% of ovarian cancers overall (Pal et al. 2005). More information is available at http://www.ncbi.nlm.nih.gov/books/NBK1247/.
Deletions or duplications are more frequent for BRCA1 than for BRCA2. The prevalence of deletions or duplications is dependent on the studied population and ranges from 0% to 11% of all BRCA2 mutations (Agata et al. 2005, Woodward et al. 2005, Casilli et al. 2006, Stadler et al. 2010).
P090-B1 probemix content: This SALSA MLPA probemix P090-B1 BRCA2 contains 51 MLPA probes with amplification products between 130 and 499 nt including 40 probes for the BRCA2 gene region and 11 reference probes that detect sequences outside this region. The identity of the genes detected by the reference probes is available online (www.mlpa.com). At least one MLPA probe is present for each exon in the BRCA2 transcript; two probes are present for exons 1 and 3, three probes are present for exons 10 and 27, and six probes are present for exon 11. One of the probes for exon 3 detects the wild type sequence of the c.156_157insAlu mutation and a reduced signal can point towards the presence of this mutation or a deletion of exon 3. In addition, there is a probe for the sequence upstream and a probe for the sequence downstream of BRCA2.
This probemix contains nine quality control fragments generating amplification products between 64 and 105 nt: four DNA Quantity Fragments (Q-fragments), three DNA Denaturation Fragments (D-fragments), and one chromosome X and one chromosome Y-specific fragment. The Q-fragments are only visible when less than 100 ng sample DNA is used. Low signal of the 88 and 96 nt fragment indicates incomplete DNA denaturation. More information on how to interpret observations on these control fragments can be found in the MLPA General Protocol.
Positive control DNA samples: In case no positive DNA sample is available in your laboratory, an artificial duplication DNA sample for this probemix (catalogue number SD024) can be ordered from MRC-Holland. This SD024 Artificial Duplication DNA will show a duplication of several probes when using the following probemixes: P002 and P087 BRCA1; P045, P090 and P077 BRCA2. The SD024 Artificial Duplication DNA is a mixture of human female genomic DNA and a titrated amount of plasmid containing selected probe target sequences. For further details, please consult the SD024 Artificial Duplication DNA product description, available online: www.mlpa.com. This SD024 is for research use only (RUO).
Sample DNA
Sample DNA developed for this product: