GSM8582156: B73_tetrad_microspore_deg_rep1; Zea mays; OTHER
Total RNA was isolated from embryos, anthers and male germ cells using TRIzol (Takara, 9109). MgCl2 (final concentration: 0.2 M) was added to prevent loss of sRNAs with low GC content (Kim et al., 2012). NaAc (pH 5.2) and glycogen were added to promote RNA precipitation. The extracted RNA was stored at −80°C in LoBind tubes (Eppendorf, 022431021). Degradome sequencing was performed as previously described (Jiang et al., 2020). In brief, total RNA was ligated to 5' RNA adapter with a MmeI recognition site at its 3′-end. After ligation, first-strand cDNA was reversely transcribed, purified with 2× AMPure XP beads (v/v) (Beckman, A63880), and amplified by PCR with 5'-biotin labeled forward primer and regular reverse primer. The amplified products were digested with ExoI and purified with 2× AMPure XP beads (v/v) (Beckman, A63880). The purified products were digested with Mmel (NEB, R0637L) and purified with Dynabeads MyOne Streptavidin C1 beads (Invitrogen, 65001). The DNA products bound to beads were used as templates for PCR with index-containing primers. The ~140-bp products were gel-purified and sequenced on an Illumina HiSeqX-Ten platform with specific sequencing primer (5'-CTACACGTTCAGAGTTCTACAGTCCGAC-3') by Annoroad Gene Technology (Beijing).
GEO
GSM8582156_r1