Pan-genome of wild and cultivated rice uncovers genetic diversity, lost and selected sequences during rice domestication

Wild relatives of crops are pivotal genetic reservoirs for enhancing stress resistance and yield of cultivated varieties. Rice (Oryza sativa L.) domestication had led to the loss of numerous beneficial traits from wild rice, especially in the AA genome group, the closest wild relatives of cultivated rice. However, existing pan-genome studies of Oryza covered only limited set of AA genome wild species or focused on non-AA genome types. In this study, by using genomic re-sequencing data of 180 wild rice varieties, we constructed pan-genomes of five AA genome types including O. barthii, O. meridionalis, O. glumaepatula, O. longistaminata, and O. rufipogon, in which each comprised 67–224 Mb of novel sequence. The detected genetic variations across these species were substantial, with SNPs ranging from 7.24 × 106 to 2.60 × 107 and InDels from 2.01 × 106 to 7.24 × 106. The results hinted a primary evolutionary gene flowing direction from O. meridionalis to O. rufipogon, and subsequently to O. sativa, reflecting a potential evolutionary trend among AA genome rice species. Notably, several nsSNPs were conserved between O. meridionalis and O. rufipogon, suggesting their importance throughout the evolutionary process. Many genes related to stress response and growth regulation were found within the lost and selected regions, among which 42 and 32 genes were differentially expressed in O. rufipogon and the cultivated variety Nipponbare under salt stress. WRKY67 gene was selected as a strong candidate for salt tolerance of rice, which was further validated by RT-qPCR, co-expression analysis, and phenotypic assessment of WRKY67 knockout mutant under salt stress. This research has filled the gap in targeted exploration of genetic resources in AA genome rice, and provided the candidate genes directly applicable for salt-tolerant rice breeding.