Northern Blot Analysis of RNA

RNA samples were mixed with an equal volume of 2× formamide loading buffer (93% formamide, 0.1× Tris/Borate/EDTA (TBE), 30 mM EDTA, 0.03% bromophenol blue, and 0.03% xylene cyanol), heated at 95 °C for 5 min and then electrophoresed on a 4% polyacrylamide/7 M urea/1× TBE denaturing gel. Then, the RNA was transferred onto a Hybond^TM-N⁺ membrane (GE Healthcare) in 1× BE at 1 A for 1–2 h and cross-linked to the membrane under UV light at 254 nm and 1200 × 100 μJ/cm². The membrane was pre-hybridized in Church buffer (0.5 M Na₂HPO₄-H₃PO₄ buffer pH 7.2, 1 mM EDTA, 7% SDS, and 1% BSA) at 35 °C for 30 min and then in Church buffer with 5′-end-labeled oligo probes^{36 (link)} (Table S3) at 35 °C overnight. Subsequently, the membrane was washed once with 2× SSC and 0.1% SDS at 50 °C for 20 min and then twice with 0.1× SSC and 0.1% SDS at 50 °C for 20 min each time. The signals on the membrane were detected with a Typhoon FLA 9500 (GE Healthcare). Probes specific to GFP mRNA³⁷ were labeled with terminal deoxynucleotidyl transferase (Thermo Scientific).