Polyploid formation is accompanied by genetic and epigenetic changes in the genome. Using DNA microarrays on flower petals, previous work has shown that there are two temporal phases of gene expression evolution in recent and natural polyploid cotton. These phases were characterized by different amounts of expression bias between the A and D homoeologs of the cotton tetraploid genome. Here RNA-seq was used to investigate gene expression in petals from diploid and allopolyploid cotton. Expression biases between A- and D-genome homoeologs were characterized for thousands of genes in tetraploid and synthetic amphihaploid (F1) petals. MethylC-seq was used to estimate levels of DNA methylation from the same tissue source used in RNA-seq. These data suggest that some of the expression bias may have been due to differential DNA methylation resulting from independent polyploidization events (the natural allopolyploid and the F1 amphi-haploid). Associations between homoeolog expression bias and DNA methylation are currently being investigated and refined.