title: IsoSeq of Podocoryna genome strain: adult male medusa
design: After being starved for four days, 200 male medusae (PCLH003) were isolated in 1.5 mL Eppendorf tubes. Following three washes in ASW and removal of excess seawater, 500 L of Trizol was added. Tissues were ground with a pellet pestle until homogenized and incubated for 12 minutes. Chloroform was added, samples were homogenized through vigorous shaking, and incubated for 3 minutes. Samples were centrifuged at 13,000 rpm for 15 minutes at 4C. The supernatant was mixed with one volume of 70% EtOH, and total RNA was extracted using the Pure Link RNA Mini Kit. RNA quality was assessed on the Agilent Tapestation, yielding 2.6 g of total RNA (RIN: 9.8). Poly(A)+ RNA was purified using oligo(dT) beads, reverse transcribed to cDNA using the Clontech SMARTer PCR cDNA Synthesis Kit, amplified by PCR, and size-selected using the BluePippin system. SMRTbell libraries were constructed with the PacBio Template Prep Kit and sequenced on the PacBio Sequel System for high-fidelity, full-length transcriptome analysis.