QTL Mapping Of Important Agronomic Traits Of O. Longistaminata And Assembly Of The Common Wild Rice Pangenome

As boosting of human population,demand for food increases swiftly.By 2030,global agricultural production may have to be doubled to meet the food needs.To effectively resolve this severe challenge,rice will play a critical role.Unfortunately,after 6,000 years of continuous selection by human and intensive breeding efforts in the last centuries,modern rice varieties that ushered in the green revolution bring about dramatic increase in rice production worldwide,but a narrowing rice genetic basis which causes not only a yield plateau in farmers'fields rice production,but also instability of rice yields under casual biotic and abiotic stresses.Thus,broadening the genetic resource pool and mapping the QTLs/genes derived from wild rice are posssibe ways to solve this plight.The main results were as follows:1.Construction of the Oryza longistaminata backcross inbred lines(BILs).In order to exploit the favorable traits hided in O.longistaminata,we crossed 93-11 as maternal parent with O.longistaminata accession.the F₁ plants were backcrossed with 93-11 for two consecutive times,after twenty generations bagged selfing,a genetically stable BC₂F₂₀ BIL population with 152 lines were successfully constructed from O.longistaminata.We resequenced the parents and BILs and compared to the reference genome“MH63”.A total of 1724138 high-quality polymorphic SNPs were genotyped and converted into 2,432 bin markers.The linkage map resulting in a total genetic distance of 789.54 c M,with the average marker interval of 1.3 c M.The results shows the average introgressed segments of each line was 22.49,with an average length of2.59Mb.The introgression segment of each line ranges from 4 to 89 and the introgressed segment length ranged from 814.05kb?22.50 Mb.In order to accurately identify the coverage of the BIL population on the O.longistaminata genome,we linear rearranged the alien genomic bins of BIL lines from chromosome 1 to chromosome 12.Alignment showed that 60 representative lines were enough to represent all of the BIL lines which covered about 99.58%of the O.longistaminata genome apart from a small gap on chromosome 6.2.Phenomics analysis the agronomic traits of the Oryza longistaminata BILs.There were a great genetic variation in yield and resistance traits.of which,the maximum grain number per panicle can reach two fold of 9311,while the maximum TGW can reach to 35.6g is 1.27 fold to 9311,while the maximum primary branch number can reach to 18 was 1.8 fold to 9311,the secondary branch number can reach to 72,which is 1.6 fold to 9311,and the maximum grain length can reach to 11.2mm,which is 1.14 fold to 9311,the maximum grain width can reach to 3.1mm,which is 1.19fold to 9311.In resistance aspect,two lines were found to be highly resistant to brown planthoppers,and 17.11%of the BILs were more resistant than 9311.The resistance to two type bacterial blight can reach to the immunity level,and 55.92%and 98.68%of the BILs have better resistance than 9311,14.47%,43.20%of the BILs have higher leaf blast resistance,and panilces blast resistance than 9311.And we had identified 10 BILs highly resistance to rice false smut,and 79.61%of the BILs have higher resistance to RFS than 9311.3.In order to further identify the useful genes for rice grain yield and resistance to disease and brown planthopper,the potential QTLs were analyzed using associated mapping based on the selected SNPs from O.longistaminata BIL lines.Results showed that except the seed setting rate,41 QTLs for seven grain yield traits,9 QTLs against brown planthopper,3 QTLs against rice false smut,1 QTL against rice leaf blast spot,2 QTLs for rice neak blast spot,7 QTLs against IV type bacterial leaf blight and 2 QTLs against V type bacterial leaf blight were detected in O.longistaminata.Of which,39QTLs for high grain yield and 22 QTLs for brown planthopper and disease resistance were the first report in Oryza species.4.Common wild rice pangenome construction.The O.rufipogon pangenome was constructed using a reference-based iterative mapping and assembly approach with 13diverse accessions that sequenced in previous study representing the native range of O.rufipogon.The assembled pangenome is 375.6 Mbp in size,27.6Mb larger than the reference genome,and contains 39,373 genes,an increase of 6.2%.Compared with the primary reference,an average of 6.09%increase for the mapping rate of raw reads of wild rice to the constructed pangenome confirms that the O.rufipogon pangenome further improves the genome completeness.Modelling the total pangenome size suggests a closed pangenome included about 39,373+/-223 genes,of which,3,2156+/-350 are core genes found in all lines.5.Large propotion of rice gene PAVs is related to disease resistance.To further test whether the presence/absence variations between O.sativa lines is associated with disease resistance or not in modern rice cultivars,we conducted a genome-wide association study of rice bacterial blight resistance based on pangenome gene PAVs markers using 357 rice accessions.Four significant PAVs for IV-biotype rice bacterial blight in Guangdong located on the new assemble contigs were identified.Six significants PAVs for V biotype rice bacterial blight was identified and one is located on chr11 which corresponding to the reported gene Xa23.Which suggest pangenome conatin a great many of resistance resource for cultivars rice improvements.6.Comparative pangenomic analysis between cultivars rice and common wild rice RGAs.To investigate the RGAs between cultivars and common wild rice,we identified the whole RGAs in the pangenomic level.Results shows 1719 RGAs was identified in the wild rice pangneome and 1614 RGAs was identified in the cultivar rice pangenome.The wild rice core genome contains 1484 RGAs greater than 1214RGAs in the cultivars rice core genome.These results showed wild rice pangenome contains more resistance resource for rice improvement,and the RGAs identified in the wild rice pangenome can laid the foundation of further resistance QTLs/gene mapping.