GSM5742256: TeloHAEC_CRISPRdel-series_A_neg_control_S29; Homo sapiens; RNA-Seq
To analyze the efficiency of enhancer deletion, genomic DNA was extracted using NucleoSpin tissue kit (Macherey-Nagel, Düren, Germany) and amplified by PCR using specific primers flanking the deletion sites using DreamTaq DNA Polymerase (EP0701, Thermo Fisher Scientific). PCR products were then analyzed by electrophoresis in 1% agarose gel.  To analyze the effects of enhancer deletion on cis target gene expression, the RNA was extracted using RNeasyPlus Micro kit (QIAGEN). RNA-Seq library preparation was carried out using QuantSeq 3' mRNA-seq Library Prep Kit FWD for Illumina (Lexogen, Vienna, Austria) according to the manufacturer's instructions. The resultant library was quantified using Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific) and its quality was checked with Bioanalyzer using High Sensitivity DNA Kit (Agilent Technologies). Individual libraries were pooled in equimolar ratio (4 nM total) and sequenced with the NextSeq 550 platform (Illumina, San Diego, California) with a single end high output kit using read length 86 cycles.