Genome sequence of Gossypium anomalum facilitates interspecific introgression breeding

Crop wild relatives are an important reservoir of natural biodiversity. However, incorporating wild geneticdiversity into breeding programs is often hampered by reproductive barriers and a lack of accurate genomicinformation. We assembled a high-quality, accurately centromere-anchored genome of Gossypium anomalum, a stress-tolerant wild cotton species. We provided a strategy to discover and transfer agronomicallyvaluable genes from wild diploid species to tetraploid cotton cultivars. With a (Gossypium hirsutum 3 G.anomalum)2 hexaploid as a bridge parent, we developed a set of 74 diploid chromosome segment substitution lines (CSSLs) of the wild cotton species G. anomalum in the G. hirsutum background. This set of CSSLsincluded 70 homozygous substitutions and four heterozygous substitutions, and it collectively containedabout 72.22% of the G. anomalum genome. Twenty-four quantitative trait loci associated with plant height,yield, and fiber qualities were detected on 15 substitution segments. Integrating the reference genome withagronomic trait evaluation of the CSSLs enabled location and cloning of two G. anomalum genes thatencode peroxiredoxin and putative callose synthase 8, respectively, conferring drought tolerance andimproving fiber strength. We have demonstrated the power of a high-quality wild-species reference genomefor identifying agronomically valuable alleles to facilitate interspecific introgression breeding in crops.