GSM9128315: RNA-seq_rice_seedling_dark to light 24h, rep2; Oryza sativa; RNA-Seq
A total of 50 ug total RNA for each of the two biological replicates was used for mRNA capture (Dynabeads mRNA DIRECT Purification Kit (Invitrogen)) following the manufacturer′ s instructions with modification. Briefly, 50 ug total RNA was diluted with H2O in 100ul volume and then denatured under 65°C for 2 min, and immediately put on the ice for exactly 2 min. Next, a total of 100ul Dynabeads were washed twice with 200ul lysis/binding buffer provided in the above kit. The washed beads were then eluted in 100ul lysis/binding buffer, which further mixed with the denatured total RNA. Then, the sample was put on the rotor to binding for 15 min at room temperature. After binding, wash buffer B was used to wash the beads for twice. 30ul H2O was used to elute the beads and immediately put on the 75°C for 2min. The eluted mRNA was collected after magnetic separation. The mRNA capture process was repeated as described above to obtain the more purified mRNA. 50ng mRNAs of each replicate were used for the library construction. All these libraries were constructed exactly following the previously published protocols