GSM6284754: High-throughput high-multiplexed second screening for splicing factors; Homo sapiens; OTHER
Qiagen RNA kit and was used For RNA extraction from isolated cardiac myocytes by following manufacturer's instructions. 1 µg total RNA was mixed with 100 fmol of mouse RASL oligos, 2 pmol of biotinylated-oligo-dT and 5 µl of streptavidin magenetic beads. Followed by one hour of annealing at 45ºC, magnetic beads were washed 3 times by washing buffer. Then ligation was achieved by adding 5U of T4 DNA ligase for 1 hour at 37ºC. After 3 times of wash, ligated oligos were eluted to water by heating at 65ºC for 5 minutes, and then PCR amplified using primers containing sequencing adaptor and barcode sequences. PCR products were purified and sequenced.