Genetic Dissection Of The Natural Variation Of Ovule Number Per Ovary In Oilseed Rape(Brassica Napus L.)

Oilseed rape（Brassica napus L.）is one of the main oil crops in the world.With the rapid increase in global population,there is an urgent need to improve oil production.Genetic improvement of seed yield of oilseed rape is necessary to meet the increasing demand for edible oil.Seed yield per plant of oilseed rape is multiply determined by three components:silique number per plant（SNPP）,seed number per silique（SNPS）and seed weight（SW）.Ovule number per ovary（ONPO）determines the upper limit and potential of seed number per silique,thus indirectly affects seed yield.Although ONPO displays a wide variation in germplasm resources,the underlying genes and regulatory mechanisms are unknown.Using an association panel and lines with extreme ONPO,the genetic dissection was performed on the natural variation of ONPO in oilseed rape.The main results were as follows:（1）The ONPO of a core association population of 327 lines showed a wide variation from 19.2 to43.8（2.28-fold）in four investigated environments,indicating a large space for genetic improvement of ovule number.Genome-wide association study identified a total of 18 association loci,which were distributed on chromosomes A01,A03,A06,A07,A09,C01,C03,C06,C07,and C09,explaining4.3-11.5%of the phenotypic variance.Three loci（q ONPO.A03-2,q ONPO.A06-2,and q ONPO.C03-2）were repeatedly detected in two environments.Comparative QTL analyses showed that all association loci were not overlapped with the reported QTLs of ovule number in oilseed rape,suggesting that they were novel.（2）Thirteen lines with more（M）or less（L）ovules were selected from this core association panel.The ONPO and the SNPS of M lines ranged from 32.8 to 40.1 and from 13.2 to 26.1,respectively.The ONPO and SNPS of L lines from 20.8 to 26.2 and from 7.8 to 18.2,respectively.The means of ONPO and SNPS of M lines（35.7 and 17.6）were significantly higher than those of L lines（24.0 and 13.1）.The difference between ONPO and SNPS of M and L line ranged from 8.6 to 25.6 and from 6.5 to 18.4,respectively,and the mean of the former（18.1）was significantly larger than that of the latter（10.9）.This indicated a large space for the genetic improvement of seed number per silique in oilseed rape.A significant positive correlation（r²=0.3811）was observed between ONPO and SNPS.From the bottom to top（representing the order of flower bud differentiation）of main inflorescence,the ONPO displayed a gradual downward trend.（3）To gain insight into the physiological processes that affect ovule number,the total leaf area,the photosynthetic rate,and the phytohormones content of the extreme lines were measured.The photosynthetic rates of M and L lines varied from 26.36 to 38.82 and from 27.47 to 38.59μmol CO₂ m^-2sec^-1,respectively,and the means of the two types of lines(32.88 and 33.97μmol CO₂m^-2sec^-1)had no significant difference.The leaf areas of M and L lines varied from 152.32 to 306.12 cm² and from144.03 to 247.34 cm²,respectively,and the mean of the two types of lines（210.40 and 199.97 cm²）also had no significant difference.Moreover,the contents of GA₄,BA,and JA were significantly higher in M lines than in L lines.In contrast,the L lines exhibited significantly higher contents of ABA and IAA than the M lines.There was no significant difference in IBA,SA,GA₁,GA₃,or zeatin content between M and L lines.（4）To identify genes involved in regulating ovule number,a comparative transcriptome analysis of M vs.L lines was performed at the ovule initiation stage.A total of 4449 differentially expressed genes（DEGs）were identified,including 2095 up-and 2354 down-regulated genes,respectively.The KEGG analysis showed that these DEGs were significantly enriched in 30 pathways including DNA,RNA,protein,signaling,transport,development,cell wall,lipid metabolism,and secondary metabolism.A total of 73 DEGs were found within the genomic regions of association loci,of which six were considered as candidates for association loci,based on their functional annotation.（5）To map the QTL of ONPO,a double haploid population was constructed from a cross between two lines that displayed significant different in ovule number.This population was subjected to genotypic analysis using the Brassica 50K SNP chip,which resulted in a high-density genetic linkage map of 2111 bins.A total of 11 QTLs were detected,which were distributed on A01,A03,A06,A07,A09,A10,C02,and C05 chromosomes,explaining 7.0-15.9%of the phenotypic variation.Furthermore,two repeatable QTLs on the A07（q ONPO.A07-1）and C02（q ONPO.C02-1）were considered as major QTLs because they displayed a relatively large effect.Three QTL（q ONPO.C0A10-1,q ONPO.C02-1,and q ONPO.C05-2）were overlapped with those reported in previous studies.However,the other 8QTLs were identified as novel and not overlapped QTLs with the previous reports.A total 27 of homologous ovule number genes were identified as candidate genes.In summary,the present study revealed the physiological,genetic and molecular basis of the natural variation of ONPO in oilseed rape,providing a theoretical basis and technical support for the further improvement of ovule number in Brassica.The following study should be focus on the breeding utilization of these ONPO QTL as well as the functional validation of their candidate genes.