BACKGROUND Germline mutations in BRCA1 and BRCA2 (BRCA1/2) have been conventionally analyzed by Sanger sequencing and multiplex ligation-dependent probe amplification (MLPA). Nowadays, next-generation sequencing (NGS) is increasingly being used in clinical genetics. The aim of this study was to evaluate the performance of NGS BRCA1/2 assays by comparing them with the conventional method. MATERIALS AND METHODS We did BRCA1/2 NGS assays of 108 breast and/or ovarian cancer patients whose BRCA1/2 mutation had been previously analyzed by Sanger sequencing and MLPA using TruSeq Custom Amplicon Design AFP2. Single-nucleotide variations (SNVs) and small insertions or deletions (InDels) were evaluated. In addition, we analyzed large genomic rearrangements (LGRs) using a coverage-based algorithm as well as a revised BRCA1/2 NGS assay (BRCAaccuTest PLUS), which additionally covered a BRCA1 promoter region. RESULTS The NGS BRCA1/2 assay detected all 20 SNVs and 21 small InDels in 56 patients. Among seven LGRs detected by MLPA, six exonic LGRs were well identified by both NGS BRCA1/2 assays. One pathogenic LGR, located on a BRCA1 promoter region, was successfully identified using revised BRCAaccuTestPLUS. CONCLUSIONS These results indicated that an NGS BRCA1/2 assay could detect most LGRs including BRCA1 promoter-region deletion as well as SNVs and small InDels. Therefore, it was applicable to clinical BRCA1/2 mutation tests.

中文翻译：

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使用下一代测序技术检测BRCA1 / 2大基因组重排，包括BRCA1启动子区域的缺失。

背景技术传统上已经通过Sanger测序和多重连接依赖性探针扩增（MLPA）分析了BRCA1和BRCA2中的种系突变（BRCA1 / 2）。如今，下一代测序（NGS）越来越多地用于临床遗传学。这项研究的目的是通过与常规方法进行比较来评估NGS BRCA1 / 2分析的性能。材料与方法我们对108例乳腺癌和/或卵巢癌患者进行了BRCA1 / 2 NGS检测，这些患者先前已使用TruSeq Custom Amplicon Design AFP2通过Sanger测序和MLPA分析了BRCA1 / 2突变。评估了单核苷酸变异（SNV）和小的插入或缺失（InDels）。此外，我们使用基于覆盖率的算法以及经过修订的BRCA1 / 2 NGS分析（BRCAaccuTest PLUS）分析了大型基因组重排（LGR），它另外覆盖了BRCA1启动子区域。结果NGS BRCA1 / 2分析检测了56例患者中的所有20例SNV和21例小型InDel。通过MLPA检测到的七个LGR中，通过两个NGS BRCA1 / 2分析都能很好地鉴定出六个外显子LGR。使用修订的BRCAaccuTestPLUS成功鉴定了位于BRCA1启动子区域的一种致病LGR。结论这些结果表明，NGS BRCA1 / 2检测可以检测到大多数LGR，包括BRCA1启动子区域缺失以及SNV和小的InDel。因此，它适用于临床BRCA1 / 2突变测试。使用修订后的BRCAaccuTestPLUS成功鉴定了位于BRCA1启动子区域上的蛋白。结论这些结果表明，NGS BRCA1 / 2检测可以检测到大多数LGR，包括BRCA1启动子区域缺失以及SNV和小的InDel。因此，它适用于临床BRCA1 / 2突变测试。使用修订后的BRCAaccuTestPLUS成功鉴定了位于BRCA1启动子区域上的蛋白。结论这些结果表明，NGS BRCA1 / 2检测可以检测到大多数LGR，包括BRCA1启动子区域缺失以及SNV和小的InDel。因此，它适用于临床BRCA1 / 2突变测试。