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Background  Previous studies exploring sequence variation in the model legume,  Medicago truncatula , relied on mapping short reads to a single reference. However, read-mapping approaches are inadequate to examine large, diverse gene families or to probe variation in repeat-rich or highly divergent genome regions.  De novo  sequencing and assembly of  M. truncatula  genomes enables near-comprehensive discovery of structural variants (SVs), analysis of rapidly evolving gene families, and ultimately, construction of a pan-genome.    Results  Genome-wide synteny based on 15  de novo M. truncatula  assemblies effectively detected different types of SVs indicating that as much as 22% of the genome is involved in large structural changes, altogether affecting 28% of gene models. A total of 63 million base pairs (Mbp) of novel sequence was discovered, expanding the reference genome space for  Medicago  by 16%. Pan-genome analysis revealed that 42% (180 Mbp) of genomic sequences is missing in one or more accession, while examination of  de novo  annotated genes identified 67% (50,700) of all ortholog groups as dispensable – estimates comparable to recent studies in rice, maize and soybean. Rapidly evolving gene families typically associated with biotic interactions and stress response were found to be enriched in the accession-specific gene pool. The nucleotide-binding site leucine-rich repeat (NBS-LRR) family, in particular, harbors the highest level of nucleotide diversity, large effect single nucleotide change, protein diversity, and presence/absence variation. However, the leucine-rich repeat (LRR) and heat shock gene families are disproportionately affected by large effect single nucleotide changes and even higher levels of copy number variation.    Conclusions  Analysis of multiple  M. truncatula  genomes illustrates the value of  de novo  assemblies to discover and describe structural variation, something that is often under-estimated when using read-mapping approaches. Comparisons among the  de novo  assemblies also indicate that different large gene families differ in the architecture of their structural variation.    Electronic supplementary material  The online version of this article (doi:10.1186/s12864-017-3654-1) contains supplementary material, which is available to authorized users.

Exploring structural variation and gene family architecture with De Novo assemblies of 15 Medicago genomes