Microbiome Analysis of AnGS Samples

High-throughput sequencing technology was used to count the microorganisms in AnGS. About 20 g samples were collected before and after the reaction and sent to Majorbio (Shanghai, China) for DNA extraction and amplicon sequencing. The first step was to isolate total genomic DNA from sludge samples using the Qiagen QIAamp Rapid DNA mini kit. In the second step, the primers 341F, 805R, Ar Ba515F and Arch806R of bacteria, archaea and methanogens were used for PCR amplification of extracted DNA. Finally, all PCR products were directly observed on an Agarose gel (2% TAE buffer) and Bandage with the AxyPrep DNA Gel Extraction Kit. Paired-end amplicon library was constructed, and Illumina MiSeq platform was used for sequencing.^{22 (link)}Cluster analysis was performed on samples using Uparse software, and the similarity between microorganisms was greater than 97%, which was defined as an OTU (operational taxonomic unit). The Simpson index, Shannon index, the Converge index, Ace index and Chao index of microorganisms were calculated using QIIME software. Statistical analysis of microbial data using R language tools.
The PICRUSt was used to calculate the metabolic function of flora and microbial gene pathway.^{23 (link)}