Abstract:?Objective The aim of the study is to explore the feasibility of ion exchange chromatography as a pre-treatment method for rat fecal samples. ?Method Rat fecal samples were collected and pretreated with three different methods: control (Con) group, centrifugation (Cen) group, and ion exchange chromatography (Chr) group. The DNA extraction quality, 16S rRNA amplicon high-throughput sequencing results, and real-time fluorescence quantitative PCR detection results of specific bacterial groups were compared between samples pretreated with different methods. ?Result The mass concentration of DNA extracted from samples of the Chr group was 1.44 times that of the Cen group and 6.49 times that of the Con group. The number of operational taxonomic unites (OTUs) in the Con group was 2 630, while the number increased to 3 220 and 3 223 in the Cen group and Chr group, respectively. α diversity and β diversity analyses indicated that the Chr group had higher microbial diversity, higher similarity and repeatability among the samples within the group. The results of real-time fluorescence quantitative PCR showed that the abundance of Bifidobacterium adolescentis was different among the three groups. Specifically, the abundance of Bifidobacterium adolescentis was the highest in the Chr group, moderate in the Cen group, and the lowest in the Con group. When the volume of bacterial suspension added increased from 100 μL to 200 μL, there was no significant difference in the abundance of Bifidobacterium adolescentis in the Con group (P>0.05), while the abundance increased to 1.32 times in the Cen group and 1.70 times in the Chr group. This indicated that ion exchange chromatography better preserved the microorganisms in rat fecal samples.Conclusion As a pretreatment method of rat fecal samples, ion exchange chromatography can improve the repeatability and sensitivity of the analysis of intestinal flora.
