title: GSM9163274: siACTL6A biol rep 3 SCC1 cells RNA-seq data; Homo sapiens; RNA-Seq
library_construction_protocol: RNA-seq: Total RNA extraction was done using Trizol/chloroform method. SCC1 cells were collected following transient ACTLA6 knockdown in 1.5 ml microcentrifuge tubes. 500 µl Trizol (Invitrogen, Cat. No: 15596026) was added to the cell pellets then phase separation was done by adding 100 µl Chloroform to each tube. The mixture was incubated for 5 minutes at RT followed by centrifugation at 12000g for 15 minutes at 4°C. Top phase was transferred carefully to a new tube and 500 µl Isopropanol was added to precipitate RNA. After 10 minutes incubation at RT, the samples were centrifuged at 12000g for 20 minutes at 4°c. Next, the supernatant was discarded and washing was done with 75 % ice-cold Ethanol twice. RNA was dried at Rt for 10 minutes and suspended with Nuclease free ultrapure water. ATAC-seq: 100,000 SCC1 cells were collected after 72 hours siRNA transfection. Nuclei preparation, tagmentation reaction and purification, PCR amplification of tagmented DNA were done according to manufacturer's protocol. RNA-seq: Libraries were made using Illumina® Stranded mRNA Prep kit (20040534) ATAC-seq: Library preparation was performed using an ATAC-seq Kit (Active Motif, Cat. No: 53150). Size distribution was analyzed using Agilent 2100 bioanalyzer.