Visual Automated Fluorescence Electrophoresis Provides Simultaneous Quality, Quantity, and Molecular Weight Spectra for Genomic DNA from Archived Neonatal Blood Spots
Visual Automated Fluorescence Electrophoresis Provides Simultaneous Quality, Quantity, and Molecular Weight Spectra for Genomic DNA from Archived Neonatal Blood Spots
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DOI：
10.1016/j.jmoldx.2013.01.003
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发表时间：
2013-05-01
影响因子：
4.1
通讯作者：
Goldman, Alicia M.
中科院分区：
文献类型：
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作者：
Klassen, Tara L.;Drabek, Janice;Goldman, Alicia M.
古特里903卡存档的干血斑（DBS）是一个独特的，但终端资源适合个人和整个人口的基因组分析。有限数量的DBS衍生的基因组DNA（gDNA）可以被全基因组扩增，产生足够的gDNA用于基因组应用，尽管成功率不同;优化从这些有限的低产量标本中分离高质量DNA是必不可少的。琼脂糖凝胶电泳和分光光度法是成熟的提取后质量控制（QC）方法，但缺乏揭示下游应用中gDNA失败的详细结构、定性或定量方面的能力。可视化自动荧光电泳（VAFE）是一种新型的QC技术，可从单微升样品中提供精确的双链DNA质量、数量和分子量。我们从瑞典新生儿库中存档超过30年的3 mm DBS中提取DNA，并在全基因组扩增之前和之后，与传统的QC方法平行，在VAFE上对DBS衍生的DNA进行了首次定量和定性分析。VAFE QC数据与PCR、测序和高密度比较基因组杂交阵列中的后续样品性能相关。我们观察到核酸数量、质量和完整性的标准化得到改善，下游基因组技术的性能也得到提高。在QC中增加VAFE测量增加了对遗传数据有效性的信心，并允许对gDNA进行具有成本效益的下游分析，以用于研究和诊断应用。
The Guthrie 903 card archived dried blood spots (DBSs) are a unique but terminal resource amenable for individual and population-wide genomic profiling. The Limited amounts of DBS-derived genomic DNA (gDNA) can be whole genome amplified, producing sufficient gDNA for genomic applications, albeit with variable success; optimizing the isolation of high-quality DNA from these finite, low-yield specimens is essential. Agarose gel electrophoresis and spectrophotometry are established postextraction quality control (QC) methods but lack the power to disclose detailed structural, qualitative, or quantitative aspects that underlie gDNA failure in downstream applications. Visual automated fluorescence electrophoresis (VAFE) is a novel QC technology that affords precise quality, quantity, and molecular weight of double-stranded DNA from a single microliter of sample. We extracted DNA from 3-mm DBSs archived in the Swedish Neonatal Repository for >30 years and performed the first quantitative and qualitative analyses of DBS-derived DNA on VAFE, before and after whole genome amplified, in parallel with traditional QC methods. The VAFE QC data were correlated with subsequent sample performance in PCR, sequencing, and high-density comparative genome hybridization array. We observed improved standardization of nucleic acid quantity, quality and integrity, and high performance in the downstream genomic technologies. Addition of VAFE measures in QC increases confidence in the validity of genetic data and allows cost-effective downstream analysis of gDNA for investigational and diagnostic applications.