Laboratory for Microbiome Sciences, RIKEN Center for Integrative Medical Sciences, 1-7-22 Suehiro-Cho Tsurumi-Ku, Yokohama, Kanagawa, 230-0045, Japan.
Graduate School of Advanced Science and Engineering, Cooperative Major in Advanced Health Science, Waseda University, 3-4-1 Ohkubo Shinjuku-Ku, Tokyo, 169-8555, Japan.
Sci Rep. 2023 Nov 11;13(1):19666. doi: 10.1038/s41598-023-46890-7.
Recovering a sufficient amount of microbial DNA from extremely low-biomass specimens, such as human skin, to investigate the community structure of the microbiome remains challenging. We developed a sampling solution containing agar to increase the abundance of recovered microbial DNA. Quantitative PCR targeting the 16S rRNA gene revealed a significant increase in the amount of microbial DNA recovered from the developed sampling solution compared with conventional solutions from extremely low-biomass skin sites such as the volar forearm and antecubital fossa. In addition, we confirmed that the developed sampling solution reduces the contamination rate of probable non-skin microbes compared to the conventional solutions, indicating that the enhanced recovery of microbial DNA was accompanied by a reduced relative abundance of contaminating microbes in the 16S rRNA gene amplicon sequencing data. In addition, agar was added to each step of the DNA extraction process, which improved the DNA extraction efficiency as a co-precipitant. Enzymatic lysis with agar yielded more microbial DNA than conventional kits, indicating that this method is effective for analyzing microbiomes of low-biomass specimens.
从极低生物量标本（如人体皮肤）中回收足够量的微生物 DNA 以研究微生物组的群落结构仍然具有挑战性。我们开发了一种含有琼脂的采样溶液，以增加回收微生物 DNA 的丰度。针对 16S rRNA 基因的定量 PCR 显示，与从极低生物量皮肤部位（如掌心和肘窝）等传统溶液相比，从开发的采样溶液中回收的微生物 DNA 量显著增加。此外，我们证实与传统溶液相比，开发的采样溶液降低了可能的非皮肤微生物的污染率，这表明微生物 DNA 的增强回收伴随着 16S rRNA 基因扩增子测序数据中污染微生物的相对丰度降低。此外，在 DNA 提取过程的每一步都添加了琼脂，作为共沉淀剂提高了 DNA 提取效率。用琼脂进行酶裂解产生的微生物 DNA 多于传统试剂盒，表明该方法对分析低生物量标本的微生物组是有效的。
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