Evaluation of multiple displacement amplification for metagenomic analysis of low biomass samples.

IF 5.1 Q1 ECOLOGY
ISME communications Pub Date : 2024-02-12 eCollection Date: 2024-01-01 DOI:10.1093/ismeco/ycae024
Melody Cabrera Ospino, Katja Engel, Santiago Ruiz-Navas, W Jeffrey Binns, Andrew C Doxey, Josh D Neufeld
{"title":"Evaluation of multiple displacement amplification for metagenomic analysis of low biomass samples.","authors":"Melody Cabrera Ospino, Katja Engel, Santiago Ruiz-Navas, W Jeffrey Binns, Andrew C Doxey, Josh D Neufeld","doi":"10.1093/ismeco/ycae024","DOIUrl":null,"url":null,"abstract":"<p><p>Combining multiple displacement amplification (MDA) with metagenomics enables the analysis of samples with extremely low DNA concentrations, making them suitable for high-throughput sequencing. Although amplification bias and nonspecific amplification have been reported from MDA-amplified samples, the impact of MDA on metagenomic datasets is not well understood. We compared three MDA methods (i.e. bulk MDA, emulsion MDA, and primase MDA) for metagenomic analysis of two DNA template concentrations (approx. 1 and 100 pg) derived from a microbial community standard \"mock community\" and two low biomass environmental samples (i.e. borehole fluid and groundwater). We assessed the impact of MDA on metagenome-based community composition, assembly quality, functional profiles, and binning. We found amplification bias against high GC content genomes but relatively low nonspecific amplification such as chimeras, artifacts, or contamination for all MDA methods. We observed MDA-associated representational bias for microbial community profiles, especially for low-input DNA and with the primase MDA method. Nevertheless, similar taxa were represented in MDA-amplified libraries to those of unamplified samples. The MDA libraries were highly fragmented, but similar functional profiles to the unamplified libraries were obtained for bulk MDA and emulsion MDA at higher DNA input and across these MDA libraries for the groundwater sample. Medium to low-quality bins were possible for the high input bulk MDA metagenomes for the most simple microbial communities, borehole fluid, and mock community. Although MDA-based amplification should be avoided, it can still reveal meaningful taxonomic and functional information from samples with extremely low DNA concentration where direct metagenomics is otherwise impossible.</p>","PeriodicalId":73516,"journal":{"name":"ISME communications","volume":null,"pages":null},"PeriodicalIF":5.1000,"publicationDate":"2024-02-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10945365/pdf/","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"ISME communications","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.1093/ismeco/ycae024","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2024/1/1 0:00:00","PubModel":"eCollection","JCR":"Q1","JCRName":"ECOLOGY","Score":null,"Total":0}
引用次数: 0

Abstract

Combining multiple displacement amplification (MDA) with metagenomics enables the analysis of samples with extremely low DNA concentrations, making them suitable for high-throughput sequencing. Although amplification bias and nonspecific amplification have been reported from MDA-amplified samples, the impact of MDA on metagenomic datasets is not well understood. We compared three MDA methods (i.e. bulk MDA, emulsion MDA, and primase MDA) for metagenomic analysis of two DNA template concentrations (approx. 1 and 100 pg) derived from a microbial community standard "mock community" and two low biomass environmental samples (i.e. borehole fluid and groundwater). We assessed the impact of MDA on metagenome-based community composition, assembly quality, functional profiles, and binning. We found amplification bias against high GC content genomes but relatively low nonspecific amplification such as chimeras, artifacts, or contamination for all MDA methods. We observed MDA-associated representational bias for microbial community profiles, especially for low-input DNA and with the primase MDA method. Nevertheless, similar taxa were represented in MDA-amplified libraries to those of unamplified samples. The MDA libraries were highly fragmented, but similar functional profiles to the unamplified libraries were obtained for bulk MDA and emulsion MDA at higher DNA input and across these MDA libraries for the groundwater sample. Medium to low-quality bins were possible for the high input bulk MDA metagenomes for the most simple microbial communities, borehole fluid, and mock community. Although MDA-based amplification should be avoided, it can still reveal meaningful taxonomic and functional information from samples with extremely low DNA concentration where direct metagenomics is otherwise impossible.

评估用于低生物量样本元基因组分析的多重位移扩增技术。
将多重置换扩增(MDA)与元基因组学相结合,可以分析DNA浓度极低的样本,使其适合高通量测序。虽然有报道称 MDA 扩增样本会产生扩增偏差和非特异性扩增,但 MDA 对元基因组数据集的影响还不甚了解。我们比较了三种 MDA 方法(即散装 MDA、乳液 MDA 和引物酶 MDA),对两种 DNA 模板浓度(约 1 pg 和 100 pg)的元基因组进行分析,这两种模板分别来自微生物群落标准 "模拟群落 "和两种低生物量环境样本(即钻孔流体和地下水)。我们评估了 MDA 对基于元基因组的群落组成、组装质量、功能特征和分选的影响。我们发现,所有 MDA 方法都存在针对高 GC 含量基因组的扩增偏差,但嵌合体、伪影或污染等非特异性扩增相对较少。在微生物群落图谱中,我们观察到了与 MDA 相关的代表性偏差,尤其是低输入 DNA 和引物酶 MDA 方法。不过,MDA 扩增文库中的分类群与未扩增样本中的分类群相似。MDA 文库高度破碎,但在较高的 DNA 输入量下,散装 MDA 和乳液 MDA 文库以及地下水样本的这些 MDA 文库都获得了与未扩增文库相似的功能图谱。对于最简单的微生物群落、井眼流体和模拟群落,高输入量的块状 MDA 元基因组可获得中低质量的分区。虽然应避免基于 MDA 的扩增,但它仍能从 DNA 浓度极低的样本中揭示有意义的分类和功能信息,否则就无法直接进行元基因组学研究。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
求助全文
约1分钟内获得全文 求助全文
来源期刊
自引率
0.00%
发文量
0
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
联系我们:info@booksci.cn Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术官方微信