title: RNA-Seq of gut microbiota:children aged 4-80 years
design: Microbic DNA was extracted by QIAamp DNA Stool Mini Kit (Qiagen) according to the manufacturer's instructions. The 16S hypervariable region of qualified DNA product was amplified with the primer (515F-805R, Invitrogen). The quantification and the purification of PCR products used respectively Quant-iTTM dsDNA Assay Kit (PicoGreen) and QIAquick PCR purification kit (Qiagen). The recovered products were amplified in a second round, and Illumina bridge PCR compatible primers were introduced. The amplified products were detected by fluorescence quantification (PicoGreen) and Aglient 2274 Bioanalyzer. After qualified, sequencing was performed using Illumina MiSeq sequencing system.