GSM6892031: Placenta_TR_rep1; Macaca fascicularis; RNA-Seq
Total RNA was extracted from the frozen placenta tissues using phenol-chloroform extraction method. In brief, 500 µl TRIZOL reagent was added into each tube containing a steel bead and placenta tissue. After crushing with griding mill, another 500 µl TRIZOL reagent was added into each tube and vertexing. After holding on ice for 10 min, 200 µl chloroform was added into each tube and hold on ice for another 10 min after vertexing. Then, the tubes were centrifuged with 12,000 rpm in 4 ℃ for 10 min. After centrifugation, the supernatant (500 µl) was mixed with 500 µl isopropanol and hold on ice for 10 min after vertexing. Next, another round of centrifugation was performed with 12,000 rpm in 4 ℃ for 10 min. After the sediments were washed with 75% ethyl alcohol, the tubes containing total RNA were dried in cabinet. The RNA libraries were finally diluted into proper concentration with RNase-free water. 1. Take a certain amount of total RNA samples, and use oligo dT beads to enrich mRNA with poly A tail;  2. mRNA molecules were fragmented into small pieces;  3. The fragmented mRNA was synthesized into first strand cDNA using random primers;  4. The second strand cDNA was synthesized;  5. The synthesized cDNA was subjected to end-repair and 3' adenylated. Adaptors were ligated to the ends of these 3' adenylated cDNA fragments;  6. PCR amplification and purification;  7. Library quality control;  8. Library circularization;  9. The library was amplified to make DNA nanoball (DNB);  10. Sequencing on DNBSEQ (DNBSEQ Technology) platform.