RNA-Seq of WC
DNA was subsequently isolated using the phenol-chloroform method. The DNA was suspended in 100 l of TE buffer, heated at 50C for 10 min, and stored at -20C. There were three replicates for control, homemade probiotic, commercial probiotic 1 and three replicates for commercial probiotic 2. Amplicon metagenomic analysis was performed by Novogene Company in China. Sequencing libraries were prepared using the TruSeq DNA PCR-Free Sample Preparation Kit (Illumina, USA) according to the manufacturer's guidelines, with index codes included. The quality of the libraries was evaluated using the Qubit@2.0 Fluorometer (Thermo Scientific) and the Agilent Bioanalyzer 2100 system. Ultimately, sequencing was carried out on an Illumina NovaSeq 6000 platform (PE250), producing (250X2) bp paired-end reads for the V4 regions of the 16S rRNA gene.