将安捷伦 2100 生物分析仪应用于下一代测序的质量控制。

IF 1.5 4区 医学 Q2 MEDICINE, LEGAL
Alina Senst PhD, Hannah Bonsiepe MSc, Sarah Kron, Iris Schulz PhD
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引用次数: 0

摘要

下一代测序(NGS)技术可对大量遗传标记进行高效、精确的基因分型,从而扩大了法医 DNA 分析的范围。然而,在复杂的样本处理和确保集合样本的等摩尔浓度方面仍存在挑战。由于最佳簇密度对测序性能至关重要,因此确定簇的数量和质量对文库制备不可或缺。在本研究中,我们调查了 Agilent 2100 生物分析仪在文库质量控制中的应用,因为法医方法的研究,尤其是针对高度降解的死后样本的研究并不多见。我们的分析包括评估总 DNA 浓度、荧光单位 (FU) 值和来自口腔拭子和腐烂尸体组织样本的纯化 DNA 文库中的适配二聚体浓度。灵敏度研究测试了从口腔拭子中提取的序列稀释液,结果显示,随着 DNA 输入浓度的降低,FU 值下降,适配器二聚体增加。安捷伦 2100 生物分析仪运行之间总 DNA 浓度和平均峰高的偏差表明数据缺乏可重复性,给精确定量带来了挑战,这在之前的研究中也有观察到。然而,对分解人类遗骸中降解样本的分析表明,有能力检测适配器二聚体的浓度,这对后续 NGS 文库制备的质量和测序的成功至关重要。因此,安捷伦 2100 生物分析仪被证明是 NGS 质量控制的重要工具。
本文章由计算机程序翻译,如有差异,请以英文原文为准。

Application of the Agilent 2100 Bioanalyzer instrument as quality control for next-generation sequencing

Application of the Agilent 2100 Bioanalyzer instrument as quality control for next-generation sequencing

Next-generation sequencing (NGS) technologies have expanded the spectrum of forensic DNA analysis by facilitating efficient and precise genotyping of a large number of genetic markers. Yet, challenges persist regarding complex sample processing and assurance of equal molar concentrations across pooled samples. Since optimal cluster density is crucial for sequencing performance, the determination of both quantity and quality is indispensable for library preparation. In this study, we investigated the application of the Agilent 2100 Bioanalyzer for library quality control, as studies for forensic approaches, particularly for highly degraded postmortem samples, are rare. Our analysis encompassed assessing total DNA concentrations, fluorescence unit (FU) values, and adapter dimer concentrations in purified DNA libraries derived from buccal swabs and tissue samples of decomposed corpses. The sensitivity study tested a serial dilution derived from buccal swabs and revealed a decrease in FU values and an increase in adapter dimers with declining DNA input concentrations. Deviations in total DNA concentrations and average peak heights between the Agilent 2100 Bioanalyzer runs indicated a lack of repeatability in data and presented challenges in accurate quantification, which was also observed in previous studies. Yet, the analysis of degraded samples from decomposed human remains has shown the ability to detect adapter dimer concentrations, which can be crucial for the quality of subsequent NGS library preparation and sequencing success. Therefore, the Agilent 2100 Bioanalyzer proves to be a valuable tool for NGS quality control.

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来源期刊
Journal of forensic sciences
Journal of forensic sciences 医学-医学:法
CiteScore
4.00
自引率
12.50%
发文量
215
审稿时长
2 months
期刊介绍: The Journal of Forensic Sciences (JFS) is the official publication of the American Academy of Forensic Sciences (AAFS). It is devoted to the publication of original investigations, observations, scholarly inquiries and reviews in various branches of the forensic sciences. These include anthropology, criminalistics, digital and multimedia sciences, engineering and applied sciences, pathology/biology, psychiatry and behavioral science, jurisprudence, odontology, questioned documents, and toxicology. Similar submissions dealing with forensic aspects of other sciences and the social sciences are also accepted, as are submissions dealing with scientifically sound emerging science disciplines. The content and/or views expressed in the JFS are not necessarily those of the AAFS, the JFS Editorial Board, the organizations with which authors are affiliated, or the publisher of JFS. All manuscript submissions are double-blind peer-reviewed.
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