Microbial Profiling from Mouse Feces

DNA was extracted from mouse faeces using the Fecal FastDNA Spin Kit (MP Biomedical, Catalogue no. 6570200) according to the manufacturer’s instructions and was then used as a template for the polymerase chain reaction amplification of the V3–V4 fragments of the bacteria. 341F and 806R were used as forward and reverse primers, respectively. After electrophoresis on a 1.5% agarose gel, the microbial genomic DNA was stained with a 4% nucleic acid dye. The DNA Gel/PCR Purification Miniprep Kit (BW-DC3511-01, BIOMIGA, San Diego, CA, USA) was used to recover and purify the PCR products in accordance with the manufacturer’s instructions. To generate DNA libraries, DNA samples were mixed at similar concentrations after measuring the concentration of the extracted DNA using a NanoDrop spectrophotometer. They were subsequently sent to a MiSeq sequencer for sequencing using a MiSeq kit (Illumina, San Diego, CA, USA), and data processing and bioinformatics analysis were carried out using the QIIME2 platform. Quality control of the downstream data was performed according to the method described by Wang et al. [36 (link)]. Visualization was performed using OmicStudio (https://www.omicstudio.cn/home, accessed on 22 July 2023).