Sensitive determination of BRAF copy number in clinical samples by pyrosequencing
Pilocytic astrocytoma is the most frequently occurring brain tumor during childhood. It is classified as grade I by the World Health Organization and may rarely evolve into higher-grade tumors. Frequent genetic abnormalities documented in astrocytomas in children are gains on chromosomal arm 7q. Dup...
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2011
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| Online Access: | http://hdl.handle.net/10725/6186 http://dx.doi.org/10.1097/PDM.0b013e3182143817 http://libraries.lau.edu.lb/research/laur/terms-of-use/articles.php http://journals.lww.com/molecularpathology/Abstract/2011/09000/Sensitive_Determination_of_BRAF_Copy_Number_in.4.aspx |
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| _version_ | 1864513466155401216 |
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| author | El Maarri, Osman |
| author2 | Setty, Prashanth Gessi, Marco Waha, Anke Waha, Andreas Torsten, Pietsch |
| author2_role | author author author author author |
| author_facet | El Maarri, Osman Setty, Prashanth Gessi, Marco Waha, Anke Waha, Andreas Torsten, Pietsch |
| author_role | author |
| dc.creator.none.fl_str_mv | El Maarri, Osman Setty, Prashanth Gessi, Marco Waha, Anke Waha, Andreas Torsten, Pietsch |
| dc.date.none.fl_str_mv | 2011 2017-09-14T09:08:23Z 2017-09-14T09:08:23Z 2017-09-14 |
| dc.identifier.none.fl_str_mv | 1533-4066 http://hdl.handle.net/10725/6186 http://dx.doi.org/10.1097/PDM.0b013e3182143817 Setty, P., Gessi, M., Waha, A., Hammes, J., El-Maarri, O., Pietsch, T., & Waha, A. (2011). Sensitive determination of BRAF copy number in clinical samples by pyrosequencing. Diagnostic Molecular Pathology, 20(3), 148-157. http://libraries.lau.edu.lb/research/laur/terms-of-use/articles.php http://journals.lww.com/molecularpathology/Abstract/2011/09000/Sensitive_Determination_of_BRAF_Copy_Number_in.4.aspx |
| dc.language.none.fl_str_mv | en |
| dc.relation.none.fl_str_mv | Diagnostic Molecular Pathology |
| dc.rights.*.fl_str_mv | info:eu-repo/semantics/openAccess |
| dc.title.none.fl_str_mv | Sensitive determination of BRAF copy number in clinical samples by pyrosequencing |
| dc.type.none.fl_str_mv | Article info:eu-repo/semantics/publishedVersion info:eu-repo/semantics/article |
| description | Pilocytic astrocytoma is the most frequently occurring brain tumor during childhood. It is classified as grade I by the World Health Organization and may rarely evolve into higher-grade tumors. Frequent genetic abnormalities documented in astrocytomas in children are gains on chromosomal arm 7q. Duplications at 7q34 lead to a fusion between genes KIAA1549 and BRAF resulting in constitutive activation of the BRAF kinase. The BRAF gene is located on chromosome 7q34 and a pseudogene has been identified on chromosome Xq13. We have developed a simple and sensitive pyrosequencing method for the determination of the BRAF copy number in clinical samples. The approach is based on the simultaneous amplification of a DNA fragment contained in exon 11 of BRAF and the respective pseudogene that is used as an internal control. Three different bases in the PCR product allow precise sequence assessment of products originating from the BRAF gene and the respective pseudogene and a calculation of gene copy numbers. After the calibration of the assay on 78 control DNA samples, 42 clinical PA samples were analyzed for variation in copy numbers by pyrosequencing and for fusion gene expression by reverse transcription-polymerase chain reaction. The results obtained from tumor DNA by the developed assay and the established reverse transcription-polymerase chain reaction assays show a high concordance. In summary, we have established a pyrosequencing-based assay allowing precise detection of BRAF copy numbers in DNA extracted from clinical samples. |
| eu_rights_str_mv | openAccess |
| format | article |
| id | LAURepo_0d1b41a77ebc9741aed3978812be0d90 |
| identifier_str_mv | 1533-4066 Setty, P., Gessi, M., Waha, A., Hammes, J., El-Maarri, O., Pietsch, T., & Waha, A. (2011). Sensitive determination of BRAF copy number in clinical samples by pyrosequencing. Diagnostic Molecular Pathology, 20(3), 148-157. |
| language_invalid_str_mv | en |
| network_acronym_str | LAURepo |
| network_name_str | Lebanese American University repository |
| oai_identifier_str | oai:laur.lau.edu.lb:10725/6186 |
| publishDate | 2011 |
| repository.mail.fl_str_mv | |
| repository.name.fl_str_mv | |
| repository_id_str | |
| spelling | Sensitive determination of BRAF copy number in clinical samples by pyrosequencingEl Maarri, OsmanSetty, PrashanthGessi, MarcoWaha, AnkeWaha, AndreasTorsten, PietschPilocytic astrocytoma is the most frequently occurring brain tumor during childhood. It is classified as grade I by the World Health Organization and may rarely evolve into higher-grade tumors. Frequent genetic abnormalities documented in astrocytomas in children are gains on chromosomal arm 7q. Duplications at 7q34 lead to a fusion between genes KIAA1549 and BRAF resulting in constitutive activation of the BRAF kinase. The BRAF gene is located on chromosome 7q34 and a pseudogene has been identified on chromosome Xq13. We have developed a simple and sensitive pyrosequencing method for the determination of the BRAF copy number in clinical samples. The approach is based on the simultaneous amplification of a DNA fragment contained in exon 11 of BRAF and the respective pseudogene that is used as an internal control. Three different bases in the PCR product allow precise sequence assessment of products originating from the BRAF gene and the respective pseudogene and a calculation of gene copy numbers. After the calibration of the assay on 78 control DNA samples, 42 clinical PA samples were analyzed for variation in copy numbers by pyrosequencing and for fusion gene expression by reverse transcription-polymerase chain reaction. The results obtained from tumor DNA by the developed assay and the established reverse transcription-polymerase chain reaction assays show a high concordance. In summary, we have established a pyrosequencing-based assay allowing precise detection of BRAF copy numbers in DNA extracted from clinical samples.PublishedN/A2017-09-14T09:08:23Z2017-09-14T09:08:23Z20112017-09-14Articleinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/article1533-4066http://hdl.handle.net/10725/6186http://dx.doi.org/10.1097/PDM.0b013e3182143817Setty, P., Gessi, M., Waha, A., Hammes, J., El-Maarri, O., Pietsch, T., & Waha, A. (2011). Sensitive determination of BRAF copy number in clinical samples by pyrosequencing. Diagnostic Molecular Pathology, 20(3), 148-157.http://libraries.lau.edu.lb/research/laur/terms-of-use/articles.phphttp://journals.lww.com/molecularpathology/Abstract/2011/09000/Sensitive_Determination_of_BRAF_Copy_Number_in.4.aspxenDiagnostic Molecular Pathologyinfo:eu-repo/semantics/openAccessoai:laur.lau.edu.lb:10725/61862021-03-19T10:00:49Z |
| spellingShingle | Sensitive determination of BRAF copy number in clinical samples by pyrosequencing El Maarri, Osman |
| status_str | publishedVersion |
| title | Sensitive determination of BRAF copy number in clinical samples by pyrosequencing |
| title_full | Sensitive determination of BRAF copy number in clinical samples by pyrosequencing |
| title_fullStr | Sensitive determination of BRAF copy number in clinical samples by pyrosequencing |
| title_full_unstemmed | Sensitive determination of BRAF copy number in clinical samples by pyrosequencing |
| title_short | Sensitive determination of BRAF copy number in clinical samples by pyrosequencing |
| title_sort | Sensitive determination of BRAF copy number in clinical samples by pyrosequencing |
| url | http://hdl.handle.net/10725/6186 http://dx.doi.org/10.1097/PDM.0b013e3182143817 http://libraries.lau.edu.lb/research/laur/terms-of-use/articles.php http://journals.lww.com/molecularpathology/Abstract/2011/09000/Sensitive_Determination_of_BRAF_Copy_Number_in.4.aspx |