Integrated transcriptomic and metabolomic analysis provides insight into the pollen development of CMS-D1 rice.

BACKGROUND: Cytoplasmic male sterility (CMS) has greatly improved the utilization of heterosis in crops due to the absence of functional male gametophyte. The newly developed sporophytic D1 type CMS (CMS-D1) rice exhibits unique characteristics compared to the well-known sporophytic CMS-WA line, making it a valuable resource for rice breeding. RESULTS: In this research, a novel CMS-D1 line named Xingye A (XYA) was established, characterized by small, transparent, and shriveled anthers. Histological and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assays conducted on anthers from XYA and its maintainer line XYB revealed that male sterility in XYA is a result of delayed degradation of tapetal cells and abnormal programmed cell death (PCD) of microspores. Transcriptome analysis of young panicles revealed that differentially expressed genes (DEGs) in XYA, compared to XYB, were significantly enriched in processes related to chromatin structure and nucleosomes during the microspore mother cell (MMC) stage. Conversely, processes associated with sporopollenin biosynthesis, pollen exine formation, chitinase activity, and pollen wall assembly were enriched during the meiosis stage. Metabolome analysis identified 176 specific differentially accumulated metabolites (DAMs) during the meiosis stage, enriched in pathways such as α-linoleic acid metabolism, flavone and flavonol biosynthesis, and linolenic acid metabolism. Integration of transcriptomic and metabolomic data underscored the jasmonic acid (JA) biosynthesis pathway was significant enriched in XYA during the meiosis stage compared to XYB. Furthermore, levels of JA, MeJA, OPC4, OPDA, and JA-Ile were all higher in XYA than in XYB at the meiosis stage. CONCLUSIONS: These findings emphasize the involvement of the JA biosynthetic pathway in pollen development in the CMS-D1 line, providing a foundation for further exploration of the molecular mechanisms involved in CMS-D1 sterility.

DNAL7, a new allele of NAL11, has major pleiotropic effects on rice architecture.

MAIN CONCLUSION: dnal7, a novel allelic variant of the OsHSP40, affects rice plant architecture and grain yield by coordinating auxins, cytokinins, and gibberellic acids. Plant height and leaf morphology are the most important traits of the ideal plant architecture (IPA), and discovering related genes is critical for breeding high-yield rice. Here, a dwarf and narrow leaf 7 (dnal7) mutant was identified from a γ-ray treated mutant population, which exhibits pleiotropic effects, including dwarfing, narrow leaves, small seeds, and low grain yield per plant compared to the wild type (WT). Histological analysis showed that the number of veins and the distance between adjacent small veins (SVs) were significantly reduced compared to the WT, indicating that DNAL7 controls leaf size by regulating the formation of veins. Map-based cloning and transgenic complementation revealed that DNAL7 is allelic to NAL11, which encodes OsHSP40, and the deletion of 2 codons in dnal7 destroyed the His-Pro-Asp (HPD) motif of OsHSP40. In addition, expression of DNAL7 in both WT and dnal7 gradually increased with the increase of temperature in the range of 27-31 °C. Heat stress significantly affected the seedling height and leaf width of the dnal7 mutant. A comparative transcriptome analysis of WT and dnal7 revealed that DNAL7 influenced multiple metabolic pathways, including plant hormone signal transduction, carbon metabolism, and biosynthesis of amino acids. Furthermore, the contents of the cytokinins in leaf blades were much higher in dnal7 than in the WT, whereas the contents of auxins were lower in dnal7. The contents of bioactive gibberellic acids (GAs) including GA1, GA3, and GA4 in shoots were decreased in dnal7. Thus, DNAL7 regulates rice plant architecture by coordinating the balance of auxins, cytokinins, and GAs. These results indicate that OsHSP40 is a pleiotropic gene, which plays an important role in improving rice yield and plant architecture.

Development of InDel markers for Oryza sativa ssp. javanica based on whole-genome resequencing.

Oryza sativa ssp. javanica rice varieties exhibit a wide variation in the phenotypes of several important agronomic traits, including grain quality, grain shape, plant architecture, disease resistance, and high adaption to an unfavorable environment, indicating a great potential for rice improvement. DNA molecular markers are basic and critical tools in genetic analysis and gene mining. However, only a few whole-genome variation analyses have been performed in Oryza sativa ssp. Javanica (tropical japonica rice), and this has hampered the utilization of such an important resource. In this study, the length of insertions/deletions variation greater larger than 10 bp from 10 Oryza sativa ssp. indica rice and 10 Oryza sativa ssp. tropical japonica rice were extracted by using the Nipponbare genome as a reference. A total of 118 primer pairs which were almost evenly distributed on each chromosome corresponding to the loci of InDels were designed by the Primer 5 program. We confirmed 85 InDel markers from 60 rice varieties, including indica and tropical japonica, by running polyacrylamide gels. The InDel markers function like SSRs in identifying hybrids, calculating genetic distance, constructing the genetic linkage map, and gene mining. The InDel markers developed in this study might help in genetic studies and to investigate the tropical japonica rice varieties.

Whole Genome Resequencing of 20 Accessions of Rice Landraces Reveals Javanica Genomic Structure Variation and Allelic Genotypes of a Grain Weight Gene TGW2.

The landraces preserved by indigenous worldwide exhibited larger variation in the phenotypes and adaption to different environments, which suggests that they comprise rich resources and can be served as a gene pool for rice improvement. Despite extensive studies on cultivated rice, the variations and relationships between landraces and modern cultivated rice remain unclear. In this study, a total of 20 varieties that include 10 Oryza javanica collected from different countries worldwide and 10 Oryza indica from China were genotyped and yielded a sum of 99.9-Gb resequencing raw data. With the genomic sequence of the japonica cultivar Nipponbare as a reference, the following genetic features of single-nucleotide polymorphism (SNP) ranged from 861,177 to 1,044,617, insertion-deletion polymorphisms (InDels) ranged from 164,018 to 211,135, and structural variation (SV) ranged from 3,313 to 4,959 were identified in Oryza javanica. Variation between the two subspecies was also determined that 584,104 SNPs, 75,351 InDels, 104,606 SNPs, and 19,872 InDels specific to Oryza indica and Oryza javanica, respectively. Furthermore, Gene Ontology (GO) and KEGG of Oryza javanica-specific SNP-related genes revealed that they participated in DNA metabolic process, DNA replication, and DNA integration. The sequence variation and candidate grain shape-related gene TGW2 were identified through Fst and sweep selective analysis. Hap4 of TGW2 is performed better than others. The whole genome sequence data and genetic variation information illustrated in this study will serve as an important gene pool for molecular breeding and facilitate genetic analysis of Oryza javanica varieties.

Genome-Wide Identification and Expansion Patterns of SULTR Gene Family in Gramineae Crops and Their Expression Profiles under Abiotic Stress in Oryza sativa.

Sulfate transporters (SULTRs), also known as H+/SO42- symporters, play a key role in sulfate transport, plant growth and stress responses. However, the evolutionary relationships and functional differentiation of SULTRs in Gramineae crops are rarely reported. Here, 111 SULTRs were retrieved from the genomes of 10 Gramineae species, including Brachypodium disachyon, Hordeum vulgare, Setaria italica, Sorghum bicolor, Zea mays, Oryza barthii, Oryza rufipogon, Oryza glabbermia and Oryza sativa (Oryza sativa ssp. indica and Oryza sativa ssp. japonica). The SULTRs were clustered into five clades based on a phylogenetic analysis. Syntheny analysis indicates that whole-genome duplication/segmental duplication and tandem duplication events were essential in the SULTRs family expansion. We further found that different clades and orthologous groups of SULTRs were under a strong purifying selective force. Expression analysis showed that rice SULTRs with high-affinity transporters are associated with the functions of sulfate uptake and transport during rice seedling development. Furthermore, using Oryza sativa ssp. indica as a model species, we found that OsiSULTR10 was significantly upregulated under salt stress, while OsiSULTR3 and OsiSULTR12 showed remarkable upregulation under high temperature, low-selenium and drought stresses. OsiSULTR3 and OsiSULTR9 were upregulated under both low-selenium and high-selenium stresses. This study illustrates the expression and evolutionary patterns of the SULTRs family in Gramineae species, which will facilitate further studies of SULTR in other Gramineae species.

Repeats in mitochondrial and chloroplast genomes characterize the ecotypes of the Oryza.

Mitochondria and chloroplast are very important organelles for organism, participating in basic life activity. Their genomes contain many repeats which can lead to a variation of genome structure. Oryza is an important genus for human beings' nutrition. Several mitochondrial and chloroplast genomes of Oryza have been sequenced, which help us to insight the distribution and evolution of the repeats in Oryza species. In this paper, we compared six mitochondrial and 13 chloroplast genomes of Oryza and found that the structures of mitochondrial genomes were more diverse than chloroplast genomes. Since repeats can change the structure of the genome, resulting in the structural diversity of the genome, we analyzed all repeats and found 31 repeats in mitochondrial and 13 repeats in chloroplast genomes. Further, we developed 21 pairs of MRS molecular markers and 12 pairs of CRS molecular markers based on mitochondrial repeats and chloroplast repeats, respectively. These molecular markers can be used to detect the repeat-mediated recombination in Oryza mitochondrial and chloroplast genomes by PCR or fluorescence quantification. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-020-01198-6.

Quantitative Trait Loci Mapping of Mineral Element Contents in Brown Rice Using Backcross Inbred Lines Derived From Oryza longistaminata.

Mineral elements play an extremely important role in human health, and are worthy of study in rice grain. Wild rice is an important gene pool for rice improvement including grain yield, disease, and pest resistance as well as mineral elements. In this study, we identified 33 quantitative trait loci (QTL) for Fe, Zn, Se, Cd, Hg, and As contents in wild rice Oryza longistaminata. Of which, 29 QTLs were the first report, and 12 QTLs were overlapped to form five clusters as qSe1/qCd1 on chromosome 1, qCd4.2/qHg4 on chromosome 4, qFe5.2/qZn5.2 on chromosome 5, qFe9/qHg9.2/qAs9.2 on chromosome 9, and qCd10/qHg10 on chromosome 10. Importantly, qSe1/qCd1, can significantly improve the Se content while reduce the Cd content, and qFe5.2/qZn5.2 can significantly improve both the Fe and Zn contents, they were delimited to an interval about 53.8 Kb and 26.2 Kb, respectively. These QTLs detected from Oryza longistaminata not only establish the basis for subsequent gene cloning to decipher the genetic mechanism of mineral element accumulation, but also provide new genetic resource for rice quality improvement.

Deciphering the Genetic Basis of Lodging Resistance in Wild Rice Oryza longistaminata.

The abuse of fertilizer results in tall rice plants that are susceptible to lodging and reduced plant yield. Hence, it is important to identify and utilize the quantitative trait loci (QTLs)/genes for lodging resistance breeding. Oryza longistaminata exhibits a strong stem and high biomass productivity, which could be a candidate gene pool for cultivars lodging resistance improvement. Here, a set of 152 BC2F20 lines derived from a cross between a cultivated line 93-11 and O. longistaminata was evaluated for lodging resistance. QTL mapping analysis combined with single-nucleotide polymorphism (SNP) marker derived from high-throughput sequencing identified 12 QTLs for stem diameter (SD), 11 QTLs for stem length (SL), and 3 QTLs for breaking strength (BS). Of which, 14 QTLs were first identified from O. longistaminata. A major QTL, qLR1, which was delimited to a region ∼80 kb on chromosome 1, increased stem diameter, stem length, and breaking strength. Another major QTL, qLR8, that was delimited in an interval ∼120 kb on chromosome 8, significantly enhanced the breaking strength. These results provide evidence that O. longistaminata can be exploited to develop lodging-resistant rice lines.

Quantitative Trait Locus Mapping of the Combining Ability for Yield-Related Traits in Wild Rice Oryza longistaminata.

In decades of hybrid rice breeding, the combining ability has been successfully used to evaluate excellent parental lines and predict heterosis. However, previous studies for the combining ability mainly focused on cultivated rice and rarely involved wild rice. In this study, for the first time, we identified 20 new quantitative trait loci (QTLs) for the combining ability in wild rice using a North Carolina II mating design. Among them, qGCA1, one of the major QTLs that can significantly improve the general combining ability of the plant height, spikelet number, and yield per plant, was delimited to an interval of about 72 kb on chromosome 1. qSCA8, another major QTL, which can significantly improve the specific combining ability of the seed-setting rate and yield per plant, was located in an interval of about 90 kb on chromosome 8. These QTLs discovered from wild rice will provide new ideas to explain the genetic mechanism of the combining ability and establish the basis for breeding of high-combining-ability rice.

QTL Mapping of Seed Vigor of Backcross Inbred Lines Derived From Oryza longistaminata Under Artificial Aging.

Seed vigor is an important character of seed quality that promotes rice to germinate rapidly from soil and developing to a strong seedling, especially in the current rice direct-sowing production system. However, previous studies for seed vigor mainly concentrate in cultivars, and less reports involving in wild rice. In this study, 152 backcross inbred lines (BILs) derived from wild rice Oryza longistaminata were genotyped with re-sequencing technology, and QTLs for seed vigor related traits under normal and artificial aging treatment were analyzed. Totally, 36 QTLs were detected, of which, eight for germination potential (GP), 10 for germination rate (GR), 9 for seedling length (SL), and 9 for root length (RL). Among these, 14 novel QTLs were identified from O. longistaminata. Of which, six QTLs were related to germination, and eight related to seedling growth under aging stress. What's more, the major QTLs q9SL1.1, q6SL1.1, and q3SL1.1 for seedling length were fallen in the same locus and fine-mapped an interval about 90 Kb. The major QTLs q9GR8.1 and q9GP8.1 related with germination were fine-mapped to an interval about 90 Kb. This work will provide us basis for breeding of high seed vigor rice in rice breeding programs and further cloning of these genes.