Truseq dna pcr free sample preparaion kit

The V4 hypervariable region of the 16S rDNA gene was ampli ed using Phusion ® High-delity PCR Master Mis with GC Buffer (New England Biolabs, US). All PCR products were visualized on agarose gels (2% in TAE buffer), and puri ed with a DNA gel extraction kit (Qiagen, Germany). Paired-End (PE) amplicon library was constructed using a TruSeq ® DNA PCR-Free Sample Preparaion Kit (Illumina, US) and quanti ed by Qubit, then sequencing was performed using the Illumina Hiseq platform (APTBIO Technology, Shanghai, China).

PCR products were mixed with same volume of 1X loading buffer (contained SYB green), electrophoresis were operated on 2% agarose gel fordetection, mixture PCR products were puri ed with AxyPrepDNA Gel Extraction Kit (AXYGEN).
Library preparation and sequencing: Following manufacturer's recommendations, sequencing libraries were generated using NEB Next®Ultra™DNA Library Prep Kit for Illumina (NEB, USA) and index codes were added. The quality of library was assessed on the Qubit@ 2.0 Fluorometer (Thermo Scienti c) and Bioanalyzer 2100 system (Agilent). Then, paired-End (PE) amplicon library was constructed using a TruSeq® DNA PCR-Free Sample Preparaion Kit (Illumina, US) and quanti ed by Qubit, then sequencing was performed using the Illumina Hiseq platform (APTBIO Technology, Shanghai, China).