用于多组学分析样品制备的单微流控装置。

IF 6.1 2区 工程技术 Q1 BIOCHEMICAL RESEARCH METHODS
Lab on a Chip Pub Date : 2025-01-17 DOI:10.1039/d4lc00919c
Ranjith Kumar Ravi Kumar, Iman Haddad, Massamba Mbacké Ndiaye, Martial Marbouty, Joëlle Vinh, Yann Verdier
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引用次数: 0

摘要

结合不同的“组学”方法,如基因组学和蛋白质组学,对微生物组的理解产生详细和完整的见解是必要的。正确的样品收集和处理以及准确的分析方法对于产生可靠的数据至关重要。我们之前开发了ChipFilter设备用于微生物样品的蛋白质组学分析。我们已经证明,该装置耦合LC-MS/MS可以成功地用于鉴定微生物蛋白质。在目前的工作中,我们已经开发了我们的工作流程来分析来自同一样品的伴随蛋白和核酸。我们在设备中使用大肠杆菌、枯草芽孢杆菌和酿酒杆菌进行裂解和蛋白水解。肽段回收后进行LC-MS/MS分析,从相同的样品中回收DNA,并通过PCR扩增成功。该工作流程进一步扩展到已知成分的复杂微生物混合物。进行了蛋白质分析,鉴定了5000多种蛋白质。对回收的DNA进行测序,其性能与未进行蛋白水解的商用试剂盒提取的DNA相当。我们的研究结果表明,ChipFilter设备适用于制备平行蛋白质组学和基因组分析的样品,这在低丰度样品的情况下尤其相关,并大大减少了采样偏差。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
A single microfluidic device for multi-omics analysis sample preparation.

Combining different "omics" approaches, such as genomics and proteomics, is necessary to generate a detailed and complete insight into microbiome comprehension. Proper sample collection and processing and accurate analytical methods are crucial in generating reliable data. We previously developed the ChipFilter device for proteomic analysis of microbial samples. We have shown that this device coupled to LC-MS/MS can successfully be used to identify microbial proteins. In the present work, we have developed our workflow to analyze concomitantly proteins and nucleic acids from the same sample. We performed lysis and proteolysis in the device using cultures of E. coli, B. subtilis, and S. cerevisiae. After peptide recovery for LC-MS/MS analysis, DNA from the same samples was recovered and successfully amplified by PCR for the 3 species. This workflow was further extended to a complex microbial mixture of known compositions. Protein analysis was carried out, enabling the identification of more than 5000 proteins. The recovered DNA was sequenced, performing comparable to DNA extracted with a commercial kit without proteolysis. Our results show that the ChipFilter device is suited to prepare samples for parallel proteomic and genomic analyses, which is particularly relevant in the case of low-abundant samples and drastically reduces sampling bias.

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来源期刊
Lab on a Chip
Lab on a Chip 工程技术-化学综合
CiteScore
11.10
自引率
8.20%
发文量
434
审稿时长
2.6 months
期刊介绍: Lab on a Chip is the premiere journal that publishes cutting-edge research in the field of miniaturization. By their very nature, microfluidic/nanofluidic/miniaturized systems are at the intersection of disciplines, spanning fundamental research to high-end application, which is reflected by the broad readership of the journal. Lab on a Chip publishes two types of papers on original research: full-length research papers and communications. Papers should demonstrate innovations, which can come from technical advancements or applications addressing pressing needs in globally important areas. The journal also publishes Comments, Reviews, and Perspectives.
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