GSM5212803: GFP-FHH rep2; Homo sapiens; RNA-Seq
Cell samples were collected with Ttizol Reagent and incubated at room temperature 5min. The homogenates was thoroughly mixed with chloroform and standby 3min at room temperature. After centrifugation of 12,000 g at 4 °C, aqueous phase was transferred and mixed with isopropanol. After centrifugation of the homogenates by 12,000 g at 4 °C, RNA sediment was washed two times with 75% ethanol and obtained the total RNA. TruSeq® RNA LT Sample Prep Kit v2 (Illumina, USA) was used to treat 2 μg RNA for each sample. Then, mRNA was purified from total RNA with RNA Purification Beads, and fragmented with Elute, Prime, Fragment Mix (EPF). First and second strand cDNAs were synthesized based on random primers. End repair was performed with End Repair Mix 2. After Adenylating 3' ends, adaptors were ligated to RNA fragments. And then, the cDNA was amplified with PCR and quantified with Qubit (Invitrogen, USA).