Mitochondrial genome sequence analysis: a custom bioinformatics pipeline substantially improves Affymetrix MitoChip v2.0 call rate and accuracy.
Mitochondrial genome sequence analysis: a custom bioinformatics pipeline substantially improves Affymetrix MitoChip v2.0 call rate and accuracy.
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线粒体基因组序列分析：定制的生物信息学流程显着提高了 Affymetrix MitoChip v2.0 的调用率和准确性。
DOI：
10.1186/1471-2105-12-402
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发表时间：
2011-10-19
影响因子：
3
通讯作者：
Falk MJ
中科院分区：
文献类型：
--
作者：
Xie HM;Perin JC;Schurr TG;Dulik MC;Zhadanov SI;Baur JA;King MP;Place E;Clarke C;Grauer M;Schug J;Santani A;Albano A;Kim C;Procaccio V;Hakonarson H;Gai X;Falk MJ
线粒体基因组序列分析对线粒体疾病的诊断评价至关重要。现有的方法在异质性检测的通量、复杂性、成本效率和灵敏度方面差异很大。Affytek MitoChip v2.0使用基因分型测序技术，可以以具有成本效益的方式完成整个人类线粒体基因组的潜在准确和高通量测序。然而，使用现有软件工具实现的相对较低的呼叫率限制了该平台在临床或研究应用中的广泛采用。在这里，我们报告了一个定制的生物信息学软件管道的设计和开发，实现了大大提高的调用率和准确性的Affytechnic MitoChip v2.0平台。我们使用这种自定义管道分析了来自24个DNA样本的MitoChip v2.0数据，这些样本代表了广泛的组织类型（18个全血，3个骨骼肌，3个细胞系），突变（5.8个内切酶对缺失和6个已知的异质性突变）和单倍型组起源。将所有结果与通过至少一种其他线粒体DNA序列分析方法获得的结果进行比较，包括桑格测序、基于变性HPLC的异源双链体分析和/或Illumina Genome Analyzer II下一代测序平台。使用我们的定制管道，所有样本的平均调用率达到99.75%。对之前通过桑格测序表征的15个样本的识别结果进行比较，发现总共有29个不一致的识别结果，这意味着碱基识别错误率估计为0.012%。我们在20个通过质量控制的样品中成功地鉴定了4个已知的异质性突变和24个其他潜在的异质性突变。使用我们优化的MitoChip过滤协议（MFP）生物信息学管道的Affytek MitoChip v2.0分析现在提供可靠、高通量和具有成本效益的全线粒体基因组测序所需的高灵敏度和准确度。这种方法为临床诊断和研究应用提供了一种可行的替代方案，可以替代传统的桑格和其他新兴的全线粒体基因组测序技术。
Mitochondrial genome sequence analysis is critical to the diagnostic evaluation of mitochondrial disease. Existing methodologies differ widely in throughput, complexity, cost efficiency, and sensitivity of heteroplasmy detection. Affymetrix MitoChip v2.0, which uses a sequencing-by-genotyping technology, allows potentially accurate and high-throughput sequencing of the entire human mitochondrial genome to be completed in a cost-effective fashion. However, the relatively low call rate achieved using existing software tools has limited the wide adoption of this platform for either clinical or research applications. Here, we report the design and development of a custom bioinformatics software pipeline that achieves a much improved call rate and accuracy for the Affymetrix MitoChip v2.0 platform. We used this custom pipeline to analyze MitoChip v2.0 data from 24 DNA samples representing a broad range of tissue types (18 whole blood, 3 skeletal muscle, 3 cell lines), mutations (a 5.8 kilobase pair deletion and 6 known heteroplasmic mutations), and haplogroup origins. All results were compared to those obtained by at least one other mitochondrial DNA sequence analysis method, including Sanger sequencing, denaturing HPLC-based heteroduplex analysis, and/or the Illumina Genome Analyzer II next generation sequencing platform. An average call rate of 99.75% was achieved across all samples with our custom pipeline. Comparison of calls for 15 samples characterized previously by Sanger sequencing revealed a total of 29 discordant calls, which translates to an estimated 0.012% for the base call error rate. We successfully identified 4 known heteroplasmic mutations and 24 other potential heteroplasmic mutations across 20 samples that passed quality control. Affymetrix MitoChip v2.0 analysis using our optimized MitoChip Filtering Protocol (MFP) bioinformatics pipeline now offers the high sensitivity and accuracy needed for reliable, high-throughput and cost-efficient whole mitochondrial genome sequencing. This approach provides a viable alternative of potential utility for both clinical diagnostic and research applications to traditional Sanger and other emerging sequencing technologies for whole mitochondrial genome analysis.
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影响因子：
5.2
作者：
Leveque, Marianne;Marlin, Sandrine;Denoyelle, Francoise
通讯作者：
Denoyelle, Francoise
影响因子：
9.8
作者：
Behar, Doron M.;Villems, Richard;Rosset, Saharon
通讯作者：
Rosset, Saharon
DOI：
10.1089/gte.2005.9.190
发表时间：
2005-09-01
期刊：
GENETIC TESTING
影响因子：
--
作者：
White, HE;Durston, VJ;Cross, NCP
通讯作者：
Cross, NCP
影响因子：
3.9
作者：
Hartmann, Anne;Thieme, Marian;Oefner, Peter J.
通讯作者：
Oefner, Peter J.
影响因子：
9.8
作者：
Li, Mingkun;Schoenberg, Anna;Stoneking, Mark
通讯作者：
Stoneking, Mark