GSM3934438: TFAMKO_RNA_clone11_CN2_6; Homo sapiens; RNA-Seq
Total RNA was extracted from confluent T75 culture flasks of TFAM CRISPR Negative Control and KO cell lines (p32) using the AllPrep DNA/RNA/Protein Kit (Qiagen #80004). Illumina's TruSeq Stranded Total RNA kit protocol was used to generate libraries.  Specifically, total RNA is converted to cDNA and size selected to 150 to 200 bp in length with 3' or 5' overhangs. End repair is performed where 3' to 5' exonuclease activity of enzymes removes 3' overhangs and the polymerase activity fills in the 5' overhangs. An 'A' base is then added to the 3' end of the blunt phosphorylated DNA fragments which prepares the DNA fragments for ligation to the sequencing adapters, which have a single 'T' base overhang at their 3' end. Ligated fragments are subsequently size selected through purification using SPRI beads and undergo PCR amplification techniques to prepare the 'libraries'.