RNA-Seq of meristem tissue from S1 Prickleless Plant 2
The tips of blackberry plants were excised in the greenhouse, immediately submerged in RNA Later (Invitrogen), placed on ice, and transported to a 4C refrigerator, where they were stored for 24h to ensure thorough penetration of the RNA Later into the tissue. Meristems were then carefully dissected under a stereomicroscope and stored at -20C until RNA extraction. RNA was extracted using the IAGEN RNeasy Plant Mini Kit (IAGEN) with buffer RLT, incorporating two apical meristems per sample. The extraction buffer was supplemented with 2.5% polyvinylpyrrolidone-40 (PVP-40) (PhytoTech Labs), and samples were incubated at 65C for 10 minutes following grinding and buffer addition. An additional wash-step was included in the protocol, and RNA was eluted in 50 L of nuclease-free water (Invitrogen). Sequencing libraries were generated using the TruSeq Stranded mRNA kit (Illumina), following the manufacturer's guidelines. Library concentration and integrity were determined using qPCR (BioRad) and Bioanalyzer (Agilent). The sequencing was performed using a ~200 cycles run on the NovaSeq 6000 to generate 100 bp paired-end reads