Human transcriptome sequencing
The RNA was broken into fragments about 300bp in length by ion disruption. RNA was used as template to synthesize the first strand of cDNA with 6-base random primers and reverse transcriptase, and the first strand cDNA was used as template for the synthesis of the second strand cDNA. After the construction of the library, PCR amplification was used to enrich the library fragments, and then the library was selected according to the fragment size. The library size was 450bp. Then, the library was inspected by Agilent 2100 Bioanalyzer, and the total concentration of the library and the effective concentration of the library were detected. Then the libraries containing different Index sequences were proportionally mixed according to the effective concentration of the library and the amount of data required for the library. The mixed library was uniformly diluted to 2nM, and single-stranded library was formed by alkali denaturation. After RNA extraction, purification and library construction, the libraries were sequenced by next-generation Sequencing (NGS) based on Illumina HiSeq Sequencing platform.