GSM1611874: vector_DGE; Homo sapiens; RNA-Seq
The total RNA was used for mRNA purification using oligo-dT magnetic beads adsorption, and then oligo-dT was used to guide reverse transcription to synthesize double-stranded cDNA.The generation of 5' ends of tags was recognized by endonuclease NlaIII, which recognizes and cuts off the CATG sites on cDNA.Then, cDNA fragments with 3' ends were purified by magnetic beads precipitation, and Illumina adapter 1 was added to their 5' ends. The junction of Illumina adapter 1 and CATG site is the recognition site of MmeI, which cuts at 17bp downstream of the CATG site, producing tags with adapter 1. After removing 3' fragments with magnetic beads precipitation, the 21 bp unique tags with adaptor 1 were purified and ligated to the adaptor 2 to form a cDNA tag library. These adapter-ligated cDNA tags were enriched after 15 cycles of linear PCR amplification. The resulting fragments were purified by PAGE gel electrophoresis. The tag library was then verified using Agilent 2100 and ABI StepOnePlus Real-Time PCR System, and sequenced using the Illumina HiSeq 2000 system according to the manufacturer’s protocols. Single read - Illumina
GEO Accession
GSM1611874