Multi-strategic RNA-seq analysis reveals a high-resolution transcriptional landscape in cotton

Currently, we integrated 4 complementary high-throughput omics techniques, including Pacbio Iso-seq for directly reading full-length isoforms, deep-depth strand-specific RNA-seq (ssRNA-seq) for quantifying expression and splicing, specialized Cap analysis gene expression (CAGE-seq) and PolyA-seq for pinpointing the transcriptional initiation and polyadenylation sites, to systematically explore the transcription landscape across 16 tissues or different organ types in G. arboreum, a diploid cotton species whose ancestor may be the donor of the A-subgenome in the allotetraploid G. hirsutum. We devised a computational pipeline named Integrative Gene Isoform Assembler (IGIA) to reconstruct accurate gene structures from improved Pacbio long reads with ssRNA-seq correction, and TSS/TES boundary information. Together with large-scale experimental validations, our data revealed a dynamic and diverse transcriptional landscape in cotton: tissue-specific gene expression, alternative usage of transcription start sites (TSSs) and alternative polyadenylation (APA), which impact many genes in various aspects including gain or loss of functional RNA motifs and conserved protein domains.