Glassing Angela, Dowd Scot E, Galandiuk Susan, Davis Brian, Jorden Jeffrey R, Chiodini Rodrick J
Department of Biological and Physical Sciences, Montana State University-Billings, Billings, Montana, United States.
Molecular Research (Mr. DNA), Shallowater, Texas, United States.
J Microbiol Methods. 2015 Dec;119:239-42. doi: 10.1016/j.mimet.2015.11.001. Epub 2015 Nov 10.
Microbial metagenomics are hindered in clinical tissue samples as a result of the large relative amount of human DNA in relation to microbial DNA acting as competitive inhibitors of downstream applications. We evaluated the LOOXSTER® Enrichment Kit to separate eukaryotic and prokaryotic DNA in submucosal intestinal tissue samples having a low microbial biomass and to determine the effects of enrichment on 16s rRNA microbiota sequencing. The enrichment kit reduced the amount of human DNA in the samples 40-70% resulting in a 3.5-fold increase in the number of 16s bacterial gene sequences detected on the Illumina MiSeq platform. This increase was accompanied by the detection of 41 additional bacterial genera and 94 tentative species. The additional bacterial taxa detected accounted for as much as 25% of the total bacterial population that significantly altered the relative prevalence and composition of the intestinal microbiota. The ability to reduce the competitive inhibition created by human DNA and the concentration of bacterial DNA may allow metagenomics to be performed on complex tissues containing a low bacterial biomass.
由于临床组织样本中人类DNA的相对含量较大，相对于微生物DNA而言，它会对下游应用起到竞争性抑制作用，因此微生物宏基因组学在临床组织样本中受到阻碍。我们评估了LOOXSTER®富集试剂盒，以分离微生物生物量较低的黏膜下肠道组织样本中的真核和原核DNA，并确定富集对16s rRNA微生物群测序的影响。该富集试剂盒使样本中的人类DNA量减少了40%-70%，导致在Illumina MiSeq平台上检测到的16s细菌基因序列数量增加了3.5倍。这一增加伴随着另外41个细菌属和94个暂定物种的检测。检测到的额外细菌分类群占总细菌种群的比例高达25%，这显著改变了肠道微生物群的相对流行率和组成。减少人类DNA产生的竞争性抑制以及细菌DNA浓度的能力，可能使宏基因组学能够在含有低细菌生物量的复杂组织上进行。
