Qubit rna assay kit in qubit 4.0 fluorometer

The concentration and integrity of total RNA extracted from PBMCs by RNeasy Mini Kit (Qiagen, Germany) were checked using the Qubit RNA Assay Kit in Qubit 4.0 Fluorometer (Life Technologies, USA) and the RNA Nano 6000 Assay Kit of the Bioanalyzer 2100 System (Agilent Technologies, USA) respectively. An rRNA-depleted cDNA library was prepared from 100 ng of RNA using Stranded Total RNA Prep Ligation with Ribo-Zero Plus kit (Illumina, USA). The final library size of 300bp was denatured and sequenced on the Illumina NextSeq 2000 platform to generate 100 bp paired-end reads.

Total RNA was extracted from PBMCs by using the RNeasy Mini Kit (Qiagen, Germany) according to the manufacturer’s instructions. The concentration and integrity of total RNA were checked using the Qubit RNA Assay Kit in Qubit 4.0 Fluorometer (Life Technologies, USA) and the RNA Nano 6000 Assay Kit of the Bioanalyzer 2100 System (Agilent Technologies, USA) respectively. A total amount of 100 ng total RNA per sample was used to prepare for the rRNA-depleted cDNA library by Stranded Total RNA Prep Ligation with Ribo-Zero Plus Kit (Illumina, USA). The final library size and quality was evaluated using an Agilent High Sensitivity DNA Kit (Agilent Technologies, CA), and the fragments were found to be between 250 and 350 bp in size. The library was sequenced using an Illumina NextSeq 2000 platform to generate 100 bp paired-end reads.