For transcriptomic analysis, the protonema tissues of WT, sog1a‐2, sog1b‐1, and sog1a‐2 sog1b‐2 were suspended in 10 ml of water and homogenized. About 0.25 OD730 of the homogenized tissues was spread on BCDAT agar medium overlaid with cellophane and cultured for 13–14 days under 16‐h white light–8‐h dark cycle at 23°C. The tissues were exposed to 60Co γ‐rays for 15 min with a dose rate of 800 Gy h−1 at TARRI. After the irradiation, the tissues were incubated under white light at 23°C for 45 min and then immediately frozen with liquid nitrogen.
About 100 mg of protonema tissues were ground with liquid nitrogen, and total RNAs were extracted using RNeasy PowerPlant Kit (QIAGEN). The RNAs were treated with RNase‐Free DNase Set (QIAGEN), followed by column purification to remove genomic DNA contamination. RNA quality was checked using an RNA 6000 Bioanalyzer Kit and RNA Nano Chip (Agilent Technologies). For library preparation, mRNA was extracted from the total RNA, and libraries were prepared using a TruSeq Stranded mRNA LT Sample Kit (Illumina). The pooled libraries were sequenced on a NextSeq 500 (Illumina) to obtain the single‐end reads of 76 bp. The obtained reads were mapped to the P. patens reference genome (v.3.3). The count data were subjected to a trimmed mean of M‐value normalization in EdgeR (McCarthy et al., 2012 (link); Robinson et al., 2010 (link)).
About 100 mg of protonema tissues were ground with liquid nitrogen, and total RNAs were extracted using RNeasy PowerPlant Kit (QIAGEN). The RNAs were treated with RNase‐Free DNase Set (QIAGEN), followed by column purification to remove genomic DNA contamination. RNA quality was checked using an RNA 6000 Bioanalyzer Kit and RNA Nano Chip (Agilent Technologies). For library preparation, mRNA was extracted from the total RNA, and libraries were prepared using a TruSeq Stranded mRNA LT Sample Kit (Illumina). The pooled libraries were sequenced on a NextSeq 500 (Illumina) to obtain the single‐end reads of 76 bp. The obtained reads were mapped to the P. patens reference genome (v.3.3). The count data were subjected to a trimmed mean of M‐value normalization in EdgeR (McCarthy et al., 2012 (link); Robinson et al., 2010 (link)).