GSM7749076: hESC RBFOX2 id.178136 (G11); Homo sapiens; RNA-Seq
RNA was extracted using TRIzol (Invitrogen #15596018) according to the manufacturer's protocol. Prior to sending RNA samples to PacBio for long read sequencing, the RNA integrity number (RIN) was calculated using the RNA 6000 Nano Kit (Agilent #5067-1511). RIN scores ranged between 8.7-9.7, indicating high quality, intact RNA. Per RNA sample, we combined 1 µg of total RNA + 2 µl 1:100 ERCC RNA Spike-in Mix #1 (Invitrogen #4456740). MAS-Seq bulk Iso-Seq libraries were generated in collaboration with PacBio using a pre-commercial protocol that implements the MAS-Seq concatenation method (Al'Khafaji et al. 2023). Briefly, cDNA molecules were concatenated into an ordered array, in this case, an 8-fold array, and sequenced on the Revio system. The concatenated array is sequenced as a HiFi read and then bioinformatically de-concatenated into segmented reads (S-reads) which represent the original cDNA molecules.