Identifying Homoeologues in Allopolyploid Species from NGS Data: Examples from Nicotiana and Glycine

Polyploidy is an important phenomenon in plant evolution, especially in angiosperms where it is estimated that 15% of all speciation events have involved polyploidy. Moreover, all seed plants are ancient polyploids, and all angiosperm genomes were shaped by one or more additional whole genome duplications. Polyploids, particularly those whose genomes retain many homoeologous genes from recent events, present challenges for any sequencing method, including next generation sequencing (NGS) transcriptomic methods, for which homoeologus gene identification can be a difficult task. One of the most common approaches is homoeolog SNP identification based on the progenitor diploid distinguishing polymorphisms (DDP). These have been used successfully in Gossypium, Tragoponon, and Glycine. Here we present examples using two alternative approaches: 1) De-novo sequence assembly and homoeolog identification by a phylogenetic approach using Nicotiana tabacum 454 data and 2) reference-guided assembly by progenitor preferential read mapping with a scoring selection for perennial Glycine species Illumina data. The Nicotiana experiment revealed that only 33% of the transcripts showed at least one polymorphism between its progenitors. N. tabacum read remapping and SNP calling over the consensus sequences revealed 3% collapsed homoeologs. N. tabacum homoeologs were identified for 5% of the consensus sequences. For the Glycine experiment around 85% of the reads mapped to the reference (G. max =soybean). 11 ± 2 % of the perennial Glycine polyploid reads mapped specifically to one of the progenitor species. Analysis with the program, Structure, supported successful homoeologous SNP separation.