Xichuan Zhai, Alex Gobbi, Witold Kot, Lukasz Krych, Dennis Sandris Nielsen, Ling Deng
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Video Abstract.</p>\",\"PeriodicalId\":18447,\"journal\":{\"name\":\"Microbiome\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":13.8000,\"publicationDate\":\"2024-10-24\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11515303/pdf/\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Microbiome\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://doi.org/10.1186/s40168-024-01935-5\",\"RegionNum\":1,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"MICROBIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Microbiome","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1186/s40168-024-01935-5","RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"MICROBIOLOGY","Score":null,"Total":0}
引用次数: 0
摘要
背景:肠道病毒组是肠道微生物组不可或缺的组成部分,在维持肠道健康方面发挥着至关重要的作用。然而,由于基因组类型(dsDNA、ssDNA、dsRNA 和 ssRNA)和拓扑结构(线性、环状或片段)固有的多样性,以及随后与当前测序文库制备方法相关的偏差,准确描绘整个肠道病毒组具有挑战性。为了克服这些问题,提高各项研究的可重复性和可比性,肠道病毒组研究亟需通用或标准化的病毒组测序文库构建方法:结果:我们将基于连接的单链文库(SSLR)制备方法重新用于病毒组研究。结果:我们将基于连接的单链文库(SSLR)制备方法重新用于病毒组研究,结果表明,SSLR方法在量化病毒DNA基因组(dsDNA和ssDNA)方面表现出卓越的效率,并且在扩增偏差最小化、覆盖率均匀性和组装病毒基因组的完整性方面优于现有的双链(Nextera)和单链(xGen、MDA + Nextera)文库制备方法。SSLR 方法可用于同时制备 DNA 和 RNA 病毒基因组文库。此外,SSLR 方法还能捕获高度修饰的噬菌体基因组,而其他文库制备方法往往会丢失这些基因组:我们介绍并改进了一种用于肠道病毒组研究的快速、简单、高效的基于连接的单链 DNA 文库制备方法。该方法与 Illumina 测序平台兼容,在 3 小时的文库制备过程中只需连接试剂,与先进的文库制备方法(xGen)相似甚至更好。我们希望这种方法能得到进一步优化、验证和广泛应用,使肠道病毒组研究更具可比性和可重复性。视频摘要。
A single-stranded based library preparation method for virome characterization.
Background: The gut virome is an integral component of the gut microbiome, playing a crucial role in maintaining gut health. However, accurately depicting the entire gut virome is challenging due to the inherent diversity of genome types (dsDNA, ssDNA, dsRNA, and ssRNA) and topologies (linear, circular, or fragments), with subsequently biases associated with current sequencing library preparation methods. To overcome these problems and improve reproducibility and comparability across studies, universal or standardized virome sequencing library construction methods are highly needed in the gut virome study.
Results: We repurposed the ligation-based single-stranded library (SSLR) preparation method for virome studies. We demonstrate that the SSLR method exhibits exceptional efficiency in quantifying viral DNA genomes (both dsDNA and ssDNA) and outperforms existing double-stranded (Nextera) and single-stranded (xGen, MDA + Nextera) library preparation approaches in terms of minimal amplification bias, evenness of coverage, and integrity of assembling viral genomes. The SSLR method can be utilized for the simultaneous library preparation of both DNA and RNA viral genomes. Furthermore, the SSLR method showed its ability to capture highly modified phage genomes, which were often lost using other library preparation approaches.
Conclusion: We introduce and improve a fast, simple, and efficient ligation-based single-stranded DNA library preparation for gut virome study. This method is compatible with Illumina sequencing platforms and only requires ligation reagents within 3-h library preparation, which is similar or even better than the advanced library preparation method (xGen). We hope this method can be further optimized, validated, and widely used to make gut virome study more comparable and reproducible. Video Abstract.
期刊介绍:
Microbiome is a journal that focuses on studies of microbiomes in humans, animals, plants, and the environment. It covers both natural and manipulated microbiomes, such as those in agriculture. The journal is interested in research that uses meta-omics approaches or novel bioinformatics tools and emphasizes the community/host interaction and structure-function relationship within the microbiome. Studies that go beyond descriptive omics surveys and include experimental or theoretical approaches will be considered for publication. The journal also encourages research that establishes cause and effect relationships and supports proposed microbiome functions. However, studies of individual microbial isolates/species without exploring their impact on the host or the complex microbiome structures and functions will not be considered for publication. Microbiome is indexed in BIOSIS, Current Contents, DOAJ, Embase, MEDLINE, PubMed, PubMed Central, and Science Citations Index Expanded.