GSM5795935: Suspended satellite cells.; Meleagris gallopavo; RNA-Seq
Cells from one confluent T75 flask for each sample were digested in 500 µl of trizol and incubated at room temperature for 5 minutes. In the next step, 100 µl of chloroform was added and the sample was shaken by vortex for 1 minutes and incubated for 5 minutes. The resulting mix was centrifuged at 12000 rcf for 20 minutes. The upper aqueous phase was transferred to a new vial on ice and cooled isopropanol (isopropyl alcohol) in the equal amount was added, vortexed and incubated on ice for 30 minutes. The vial was centrifuged in a refrigerated centrifuge for 20 minutes. When the RNA pellet was visible supernatants were removed and in order to remove isopropanol it was washed twice with 75% ethanol. Ethanol was removed and the pellet was dried in room temperature for 30 minutes. Afterwards, the dried pellet was dissolved in 25 µl DEPC water and kept in -80 ° C. The NanoDrop ND2000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA USA) was used to assess RNA quantity and purity.  Five hundred nanograms of total RNA from each sample were submitted to the North Carolina State University Genomics Research Laboratory for library preparation and sequencing on the Illumina NovoSeq 6000 (Illumina Inc, San Diego CA, USA). Each sample generated 35-40 million sequence reads.