Abstract Allopolyploids contain two or more sets of subgenomes. To establish a compatible relationship between subgenomes, a series of gene expression changes occurred in allopolyploids. What evolutionary changes of transcripts have taken place in Brassica napus during the early establishment and subsequent evolution was a fascinating scientific question. Here, we study this issue using a set of materials (natural, resynthesized B. napus and their progenitors/parents) by long-read RNA sequencing technology. The results showed that more genes were up-regulated in resynthesized B. napus compared with its two parents, and more up-regulated expressed genes were observed in natural B. napus compared with resynthesized B. napus. The presence of up-regulation genes in organism may help it adapt to the influence of “genomic shock” and cope with natural environment. Isoforms are produced from precursor mRNAs by alternative splicing (AS) events, and more than 60% of novel isoforms were identified in all materials, which could improve the reference genome information of B. napus. We found that the isoform numbers, the number of genes potentially involved in AS and alternative polyadenylation increased in B. napus after evolution, which may involve in the adaptation of plants to natural environment. In addition, all identified isoforms were functional annotated by searching 7 databases. In general, this study could improve our overall understanding of the full-length transcriptome of B. napus, and help us recognize the significant gene expression changes and isoform abundance changes occurred in allopolyploid B. napus during evolution.