Location of Balanced Chromosome-Translocation Breakpoints by Long-Read Sequencing on the Oxford Nanopore Platform
Location of Balanced Chromosome-Translocation Breakpoints by Long-Read Sequencing on the Oxford Nanopore Platform
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在 Oxford Nanopore 平台上通过长读长测序定位平衡染色体易位断点
DOI：
10.3389/fgene.2019.01313
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发表时间：
2020-01-14
影响因子：
3.7
通讯作者：
Lin, Ge
中科院分区：
文献类型：
--
作者：
Hu, Liang;Liang, Fan;Lin, Ge
基因组结构变异，包括易位、倒位、插入、缺失和重复，很难通过传统基因组技术可靠地检测到。特别是，平衡易位和倒位既不能通过微阵列来识别，因为它们不会改变染色体拷贝数，也不能通过短读长测序来识别，因为短读长无法针对重复基因组区域进行定位。断点的精确定位对于探索平衡易位或倒位患者的遗传原因至关重要。长读长测序技术可以更直接、高效、准确的方式检测这些结构变异。在这里，我们使用 Oxford Nanopore GridION 测序仪进行全基因组长读长测序，以检测 6 个平衡染色体易位载体和 1 个倒位载体中的断点。结果显示，所有断点与核型结果一致，仅10x覆盖度相似。聚合酶链式反应 (PCR) 和桑格测序证实了 14 个断点中的 8 个；然而，其他断点基因座略有遗漏，因为它们要么位于高度重复区域，要么位于着丝粒周围区域。一些断点中断了正常的基因结构，而在其他情况下，在断点旁边发现了微缺失/插入。我们还检测到断点区域周围的单倍型。我们的结果表明，长读长全基因组测序是精确定位易位断点和提供单倍型信息的理想策略，这对于医学遗传学和植入前基因检测至关重要。
Genomic structural variants, including translocations, inversions, insertions, deletions, and duplications, are challenging to be reliably detected by traditional genomic technologies. In particular, balanced translocations and inversions can neither be identified by microarrays since they do not alter chromosome copy numbers, nor by short-read sequencing because of the unmappability of short reads against repetitive genomic regions. The precise localization of breakpoints is vital for exploring genetic causes in patients with balanced translocations or inversions. Long-read sequencing techniques may detect these structural variants in a more direct, efficient, and accurate manner. Here, we performed whole-genome, long-read sequencing using the Oxford Nanopore GridION sequencer to detect breakpoints in six balanced chromosome translocation carriers and one inversion carrier. The results showed that all the breakpoints were consistent with the karyotype results with only similar to 10x coverage. Polymerase chain reaction (PCR) and Sanger sequencing confirmed 8 out of 14 breakpoints; however, other breakpoint loci were slightly missed since they were either in highly repetitive regions or pericentromeric regions. Some of the breakpoints interrupted normal gene structure, and in other cases, micro-deletions/insertions were found just next to the breakpoints. We also detected haplotypes around the breakpoint regions. Our results suggest that long-read, whole-genome sequencing is an ideal strategy for precisely localizing translocation breakpoints and providing haplotype information, which is essential for medical genetics and preimplantation genetic testing.