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1.
PLoS Biol ; 17(1): e3000107, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30629605

RESUMO

Current molecular biology laboratories rely heavily on the purification and manipulation of nucleic acids. Yet, commonly used centrifuge- and column-based protocols require specialised equipment, often use toxic reagents, and are not economically scalable or practical to use in a high-throughput manner. Although it has been known for some time that magnetic beads can provide an elegant answer to these issues, the development of open-source protocols based on beads has been limited. In this article, we provide step-by-step instructions for an easy synthesis of functionalised magnetic beads, and detailed protocols for their use in the high-throughput purification of plasmids, genomic DNA, RNA and total nucleic acid (TNA) from a range of bacterial, animal, plant, environmental and synthetic sources. We also provide a bead-based protocol for bisulfite conversion and size selection of DNA and RNA fragments. Comparison to other methods highlights the capability, versatility, and extreme cost-effectiveness of using magnetic beads. These open-source protocols and the associated webpage (https://bomb.bio) can serve as a platform for further protocol customisation and community engagement.


Assuntos
Sequenciamento de Nucleotídeos em Larga Escala/métodos , Ácidos Nucleicos/isolamento & purificação , Animais , DNA/isolamento & purificação , Humanos , Campos Magnéticos , Microesferas , RNA/isolamento & purificação
2.
Cell Rep ; 39(13): 111017, 2022 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-35767948

RESUMO

Aging is characterized by a chronic low-grade inflammation known as inflammaging in multiple tissues, representing a risk factor for age-related diseases. Dietary restriction (DR) is the best-known non-invasive method to ameliorate aging in many organisms. However, the molecular mechanism and the signaling pathways that drive inflammaging across different tissues and how they are modulated by DR are not yet understood. Here we identify a multi-tissue gene network regulating inflammaging. This network is characterized by chromatin opening and upregulation in the transcription of innate immune system receptors and by activation of interferon signaling through interferon regulatory factors, inflammatory cytokines, and Stat1-mediated transcription. DR ameliorates aging-induced alterations of chromatin accessibility and RNA transcription of the inflammaging gene network while failing to rescue those alterations on the rest of the genome. Our results present a comprehensive understanding of the molecular network regulating inflammation in aging and DR and provide anti-inflammaging therapeutic targets.


Assuntos
Envelhecimento , Inflamação , Envelhecimento/fisiologia , Cromatina , Humanos , Imunidade Inata , Inflamação/metabolismo , Interferons/metabolismo , Receptores Imunológicos/metabolismo , Regulação para Cima
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