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Background A pangenome aims to capture the complete genetic diversity within a species and reduce bias in genetic analysis inherent in using a single reference genome. However, the current linear format of most plant pangenomes limits the presentation of position information for novel sequences. Graph pangenomes have been developed to overcome this limitation. However, bioinformatics analysis tools for graph format genomes are lacking. Results To overcome this problem, we develop a novel strategy for pangenome construction and a downstream pangenome analysis pipeline (PSVCP) that captures genetic variants’ position information while maintaining a linearized layout. Using PSVCP, we construct a high-quality rice pangenome using 12 representative rice genomes and analyze an international rice panel with 413 diverse accessions using the pangenome as the reference. We show that PSVCP successfully identifies causal structural variations for rice grain weight and plant height. Our results provide insights into rice population structure and genomic diversity. We characterize a new locus ( qPH8-1 ) associated with plant height on chromosome 8 undetected by the SNP-based genome-wide association study (GWAS). Conclusions Our results demonstrate that the pangenome constructed by our pipeline combined with a presence and absence variation-based GWAS can provide additional power for genomic and genetic analysis. The pangenome constructed in this study and the associated genome sequence and genetic variants data provide valuable genomic resources for rice genomics research and improvement in future.

Global food security depends heavily on a few staple crops, while orphan crops, despite being less studied, offer the potential benefits of environmental adaptation and enhanced nutritional traits, especially in a changing climate. Major crops have benefited from genomics-based breeding, initially using single genomes and later pangenomes. Recent advances in DNA sequencing have enabled pangenome construction for several orphan crops, offering a more comprehensive understanding of genetic diversity. Orphan crop research has now entered the pangenomics era and applying these pangenomes with advanced selection methods and genome editing technologies can transform these neglected species into crops of broader agricultural significance. A single reference genome is not enough to harness the full genetic variation of a species. Here, the authors review pangenomics principles, construction and applications for orphan crop improvement, and discuss the possible ways to support pangenomics-driven molecular breeding.

BackgroundCadmium (Cd) accumulation in rice followed by transfer to the food chain causes severe health problems in humans. Breeding of low Cd accumulation varieties is one of the most economical ways to solve the problem. However, information on the identity of rice germplasm with low Cd accumulation is limited, particularly in indica, and the genetic basis of Cd accumulation in rice is not well understood.ResultsScreening of 312 diverse rice accessions revealed that the grain Cd concentrations of these rice accessions ranged from 0.12 to 1.23 mg/kg, with 24 accessions less than 0.20 mg/kg. Three of the 24 accessions belong to indica. Japonica accumulated significantly less Cd than indica (p < 0.001), while tropical japonica accumulated significantly less Cd than temperate japonica (p < 0.01). GWAS in all accessions identified 14 QTLs for Cd accumulation, with 7 identified in indica and 7 identified in japonica subpopulations. No common QTL was identified between indica and japonica. The previously identified genes (OsHMA3, OsNRAMP1, and OsNRAMP5) from japonica were colocalized with QTLs identified in japonica instead of indica. Expression analysis of OsNRAMP2, the candidate gene of the novel QTL (qCd3–2) identified in the present study, demonstrated that OsNRAMP2 was mainly induced in the shoots of high Cd accumulation accessions after Cd treatment. Four amino acid differences were found in the open reading frame of OsNRAMP2 between high and low Cd accumulation accessions. The allele from low Cd accumulation accessions significantly increased the Cd sensitivity and accumulation in yeast. Subcellular localization analysis demonstrated OsNRAMP2 expressed in the tonoplast of rice protoplast.ConclusionThe results suggest that grain Cd concentrations are significantly different among subgroups, with Cd concentrations decreasing from indica to temperate japonica to tropical japonica. However, considerable variations exist within subgroups. The fact that no common QTL was identified between indica and japonica implies that there is a different genetic basis for determining Cd accumulation between indica and japonica, or that some QTLs for Cd accumulation in rice are subspecies-specific. Through further integrated analysis, it is speculated that OsNRAMP2 could be a novel functional gene associated with Cd accumulation in rice.

Background Southern rice black-streaked dwarf virus (SRBSDV), transmitted by the white-backed planthopper (WBPH), causes severe yield losses in rice across Asia. However, elite resistant germplasms and molecular defense mechanisms remain elusive, hindering breeding efforts. Results Screening of 195 international diverse rice accessions identified a highly resistant (up to 0% disease incidence) variety (R91), exhibiting dual resistance to SRBSDV and rice black-streaked dwarf virus (RBSDV). Multi-omics analysis revealed a rapid defense activation in R91, with an increase in Jasmonic Acid (JA) and Jasmonic Acid-Isoleucine (JA-Ile) by 5 days post-inoculation (dpi), along with upregulation of more than 2,000 defense genes. In contract, susceptible line showed declining JA and JA-Ile level along with suppressed defense responses. Time-ordered co-expression networks pinpointed that OsJAR2 (encoding a JA-Ile synthase, LOC_Os01g12160 ) may act as a hub of resistance molecular network. Genome wide association study (GWAS) identified a novel SRBSDV resistance quantitative trait locus ( qSRBSDV1-1 ) co-localizing with OsJAR2 , and haplotype analysis validated OsJAR2 as the candidate causal resistance gene, further providing genetic evidence for its role in SRBSDV defense. Conclusions Our study identifies a highly SRBSDV-resistant rice germplasm, offering valuable genetic resource for both resistance research and breeding programs. We demonstrate that rapid JA biosynthesis activation and coordinated defense gene expression form the molecular basis of resistance in this accession. Crucially, we pinpoint OsJAR2 as a novel functional resistance gene, with its associated resistant haplotype serving as a robust molecular marker for marker-assisted selection (MAS). These findings advance our understanding of SRBSDV resistance mechanisms and provide a genetic toolkit for developing elite, disease-resistant rice varieties.

The mechanical properties, capillary water absorption, drying shrinkage, and morphology of high-volume fly ash mortar were investigated. The mortar contained 0~2.5 wt. % nano-silicon dioxide (nano-SiO2, NS) and 0~1.5 vol.% polyvinyl alcohol (PVA) fiber, with fly ash (FA) replacing 50% of cement by weight. The experimental results demonstrated that the synergistic incorporation of NS and PVA fiber significantly improved the mortar’s mechanical performance. At 7 days of age, the flexural and compressive strength of mortar containing 1.5% NS and 1.0% PVA fiber increased by 105.8% and 25.1%, respectively, compared to the control mortar (without NS or PVA fiber). Moreover, NS and PVA fiber significantly reduced the capillary absorption rate and drying shrinkage. The composite addition of 2.0% NS and 1.0% PVA fiber led to a notable reduction in drying shrinkage: at 7, 14, 28, 90, and 180 days, the drying shrinkage decreased by 38.3%, 33.3%, 30.0%, 31.6%, and 31.4%, respectively, relative to the control mortar. The scanning electron microscopy and mercury intrusion porosimetry results indicated that NS and PVA fibers effectively improved the micropore structure of the mortar.

As one of the most crucial components, the catalyst layer (CL) plays a critical role in the performance of anion exchange membrane fuel cells (AEMFCs). However, the effect of the structural evolution of ionomer binder on the micromorphology and catalytic activity of CL is yet to be clarified. In this study, pyrrolidinum and quaternary ammonium cations are attached to the polyphenylene oxide (PPO) backbone through flexible spacer units (five, seven, or nine carbon atoms) with different terminal alkyl groups. The Van der Waals force and electrostatic repulsion between the ionomer binder and catalyst are regulated through the flexible spacer units and terminal alkyl groups to alleviate the agglomeration of catalyst particles and acquire a high catalytic activity. To evaluate the electrochemical stability of the cationic groups, the alkaline stability of the ionomer binder is tested under a constant voltage to simulate the true operational environment of the fuel cells. The results reveal that the degradation of the cation groups of ionomer binder is accelerated under a constant voltage condition. This phenomenon in neglect earlier, may serve as a useful reference for the synthesis and performance enhancement of ionomer binders. Reasonable interaction regulation between the catalyst and ionomer binder forms a high performance catalyst layer. Here, the multiple interactions, including the Van der Waals force and electrostatic repulsion between the ionomer binder and catalyst, are regulated through the flexible spacer units and terminal alkyl groups, which alleviate the agglomeration of catalyst particles and acquire a high catalytic activity.

Understanding the evolutionary forces in speciation is a central goal in evolutionary biology. Asian cultivated rice has two subspecies, indica and japonica , but the underlying mechanism of the partial reproductive isolation between them remains obscure. Here we show a presence-absence variation (PAV) at the Se locus functions as an indica-japonica reproductive barrier by causing hybrid sterility (HS) in indica-japonica crosses. The locus comprises two adjacent genes: ORF3 encodes a sporophytic pollen killer, whereas ORF4 protects pollen in a gametophytic manner. In F 1 of indica-japonica crosses, pollen with the japonica haplotype, which lacks the sequence containing the protective ORF4 , is aborted due to the pollen-killing effect of ORF3 from indica . Evolutionary analysis suggests ORF3 is a gene associated with the Asian cultivated rice species complex, and the PAV has contributed to the reproductive isolation between the two subspecies of Asian cultivated rice. Our analyses provide perspectives on rice inter-subspecies post-zygotic isolation, and will promote efforts to overcome reproductive barriers in indica-japonica hybrid rice breeding. The mechanisms of the partial reproductive isolation between the two subspecies of rice remains obscure. Here, the authors show that the two adjacent genes form a killer-protector system to induce hybrid male sterility and reproductive isolation between indica and japonica .

Background WRKY proteins are one of the largest gene families and are well-known for their regulatory roles in many aspects of plant development, including plant response to both biotic and abiotic stresses. Although the roles of WRKY proteins in leaf blast resistance have been well-documented in rice, their functions in panicle blast, the most destructive type of blast disease, are still largely unknown. Results Here, we identified that the transcription of OsWRKY67 was strongly activated by leaf and panicle blast infection. OsWRKY67 is ubiquitously expressed and sub-localized in the nucleus. Rice plants overexpressing OsWRKY67 showed quantitatively enhanced resistance to leaf blast, panicle blast and bacterial blight. In contrast, silencing of OsWRKY67 increased the susceptibility to blast and bacterial blight diseases. RNA-seq analysis indicated that OsWRKY67 induces the transcription of a set of defense-related genes including the ones involved in the salicylic acid (SA)-dependent pathway. Consistent with this, the OsWRKY67- overexpressing plants accumulated higher amounts of endogenous SA, whereas lower endogenous SA levels were observed in OsWRKY67- silenced plants relative to wild-type Nipponbare plants before and after pathogen attack. Moreover, we also observed that OsWRKY67 directly binds to the promoters of PR1a and PR10 to activate their expression. Conclusions These results together suggest the positive role of OsWRKY67 in regulating rice responses to leaf blast, panicle blast and bacterial blight disease. Furthermore, conferring resistance to two major diseases makes it a good target of molecular breeding for crop improvement in rice.

BackgroundGrain shape is a key trait in rice breeding. Although many QTLs and genes of grain shape have been identified, how different combinations of alleles of these genes affect grain shape is largely unknown. It is important to understand the effects of grain shape gene combinations for breeding by design. In the present study, we performed genetic dissection of the grain shapes in Guangdong Simiao varieties, a popular kind of rice in South China, to identify the effective alleles and their combination for breeding.ResultsWe selected two hundred nineteen indica accessions with diverse grain shapes and fifty-two Guangdong Simiao varieties with long and slender grain shapes for genome-wide selection analysis. The results showed that four (GS3, GS5, GW5 and GL7) of the twenty grain shape genes fall into the regions selected for in Guangdong Simiao varieties. Allele analysis and frequency distribution of these four genes showed that GS3allele3 and GW5allele2 accounted for 96.2%, and GL7allele2 and GS5allele2 accounted for 76.9% and 74.5% of the Simiao varieties, respectively. Further analysis of the allelic combinations showed that 30 allelic combinations were identified in the whole panel, with 28 allelic combinations found in the international indica accessions and 6 allelic combinations found in Guangdong Simiao varieties. There were mainly three combinations (combinations 17, 18 and 19) in the Guangdong Simiao varieties, with combination 19 (GS3allele3 + GW5allele2 + GL7allele2 + GS5allele2) having the highest percentage (51.9%). All three combinations carried GS3allele3 + GW5allele2, while combinations 17 (GL7allele1) and 19 (GL7allele2) showed significant differences in both grain length and length/width ratio due to differences in GL7 alleles. Pedigree analysis of Guang8B, the maintainer of the first released Simiao male sterile line Guang8A, showed that the parent lines and Guang8B carried GS3allele3 + GW5allele2 + GS5allele2, while the GL7 allele differed, resulting in significant differences in grain size.ConclusionThe results suggest that specific alleles of GS3, GS5, GW5 and GL7 are the key grain shape genes used in the Guangdong Simiao varieties and selected for grain shape improvement. Combination 19 is the predominant allelic combination in the Guangdong Simiao varieties. Our current study is the first to dissect the genetics of grain shape in Guangdong Simiao varieties, and the results will facilitate molecular breeding of Guangdong Simiao varieties.

BackgroundBacterial blight (BB), caused by Xanthomonas oryzae pv. oryzae (Xoo), is one of the most devastating diseases in rice all over the world. Due to the diversity and rapid evolution of Xoo, identification and use of the non-race specific quantitative resistance QTLs has been considered the preferred strategy for effective control of this disease. Although numerous QTLs for BB resistance have been identified, they haven’t been effectively used for improvement of BB resistance in rice due to their small effects and lack of knowledge on the function of genes underlying the QTLs.ResultsIn the present study, a genome-wide association study of BB resistance was performed in a rice core collection from South China. A total of 17 QTLs were identified to be associated with BB resistance. Among them, 13 QTLs were newly identified in the present study and the other 4 QTLs were co-localized with the previously reported QTLs or Xa genes that confer qualitative resistance to Xoo strains. Particularly, the qBBR11–4 on chromosome 11 explained the largest phenotypic variation in this study and was co-localized with the previously identified QTLs for BB and bacterial leaf streak (BLS) resistance against diverse strains in three studies, suggesting its broad-spectrum resistance and potential value in rice breeding. Through combined analysis of differential expression and annotations of the predicted genes within qBBR11–4 between two sets of rice accessions selected based on haplotypes and disease phenotypes, we identified the transcription factor OsMYB21 as the candidate gene for qBBR11–4. The OsMYB21 overexpressing plants exhibited decreased resistance to bacterial blight, accompanied with down-regulation of several defense-related genes compared with the wild-type plants.ConclusionThe results suggest that OsMYB21 negatively regulates bacterial blight resistance in rice, and this gene can be a promising target in rice breeding by using the gene editing method. In addition, the potential candidate genes for the 13 novel QTLs for BB resistance were also analyzed in this study, providing a new source for cloning of genes associated with BB resistance and molecular breeding in rice.