title: GSM8865671: STARR-seq_EE_input_rep2; Homo sapiens; OTHER
library_name: GSM8865671
library_construction_protocol: gDNA: After 24 h, cells from each replicate were pelleted, and gDNA was extracted using the Flexigene protocol (Qiagen) in a final volume of 50 µL. RNA: Total RNA was isolated using the RNeasy kit (Qiagen), followed by mRNA enrichment with Dynabeads (Invitrogen). cDNA was synthesized with Superscript III (Thermo Fisher) and a reverse-transcription primer, treated with RNase H, purified, and quantified. cDNA was then PCR-amplified (KAPA HiFi) prior to library indexing. A custom oligo pool of 11,999 sequences (Twist Bioscience) was synthesized with short 15 bp adapters corresponding to TruSeq indices. These oligos were PCR-amplified to add homology arms, then cloned into a STARR-seq vector (digested with AgeI-HF, SalI-HF, NotI-HF) via Gibson Assembly. The assembly was transformed into MegaX DH10B T1R cells (Thermo Fisher), and colonies were expanded in Terrific Broth. The resulting plasmid library (Exonhancer) was purified using midiprep kits, yielding ~1.75 mg of DNA.
identifiers.namespace: GEO
identifiers.id: GSM8865671_r1