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Background   Setaria italica  is the second-most widely planted species of millets in the world and an important model grain crop for the research of C4 photosynthesis and abiotic stress tolerance. Through three genomes assembly and annotation efforts, all genomes were based on next generation sequencing technology, which limited the genome continuity.    Results  Here we report a high-quality whole-genome of new cultivar Huagu11, using single-molecule real-time sequencing and High-throughput chromosome conformation capture (Hi-C) mapping technologies. The total assembly size of the Huagu11 genome was 408.37 Mb with a scaffold N50 size of 45.89 Mb. Compared with the other three reported millet genomes based on the next generation sequencing technology, the Huagu11 genome had the highest genomic continuity. Intraspecies comparison showed about 94.97 and 94.66% of the Yugu1 and Huagu11 genomes, respectively, were able to be aligned as one-to-one blocks with four chromosome inversion. The Huagu11 genome contained approximately 19.43 Mb Presence/absence Variation (PAV) with 627 protein-coding transcripts, while Yugu1 genomes had 20.53 Mb PAV sequences encoding 737 proteins. Overall, 969,596 Single-nucleotide polymorphism (SNPs) and 156,282 insertion-deletion (InDels) were identified between these two genomes. The genome comparison between Huagu11 and Yugu1 should reflect the genetic identity and variation between the cultivars of foxtail millet to a certain extent. The Ser-626-Aln substitution in acetohydroxy acid synthase ( AHAS ) was found to be relative to the imazethapyr tolerance in Huagu11.    Conclusions  A new improved high-quality reference genome sequence of  Setaria italica  was assembled, and intraspecies genome comparison determined the genetic identity and variation between the cultivars of foxtail millet. Based on the genome sequence, it was inferred that the Ser-626-Aln substitution in  AHAS  was responsible for the imazethapyr tolerance in Huagu11. The new improved reference genome of  Setaria italica  will promote the genic and genomic studies of this species and be beneficial for cultivar improvement.    Supplementary Information  The online version contains supplementary material available at 10.1186/s12870-021-03003-8.

bmc plant biology

De novo genome assembly of a foxtail millet cultivar Huagu11 uncovered the genetic difference to the cultivar Yugu1, and the genetic mechanism of imazethapyr tolerance