Mouse feces bacterial flora

To further identify the specific bacterial groups related to the anti-aging of UC-MSCs, we employed LEfSe to compare the intestinal flora across the Control, the D-gal, and the UC-MSCs groups. Genomic DNA was extracted from mouse feces using DNeasy PowerSoil Pro Kit. The quantity and quality of genomic DNA were determined by NanoDrop2000 spectrophotometer and agarose gel electrophoresis, respectively. PCR primers were designed based on the conserved regions of the sequence, and specific barcode sequences were incorporated corresponding to groups. PCR amplification of the 16 S V3-V4 region was followed by fluorescence quantification of the recovered PCR products. Based on the fluorescence quantification results, the samples were combined in corresponding proportions that were required for the sequencing volume. The sequencing library was constructed using NEB's NEXTFLEX Rapid DNA-Seq Kit, and subsequent high-throughput sequencing was carried out. The Miseq PE300/ NovaSeq PE300 platform was used to conduct all procedures, sequence analysis, and DNA extraction.