Integrated meta-analysis and transcriptomics pinpoint genomic loci and novel candidate genes associated with submergence tolerance in rice.

BACKGROUND: Due to rising costs, water shortages, and labour shortages, farmers across the globe now prefer a direct seeding approach. However, submergence stress remains a major bottleneck limiting the success of this approach in rice cultivation. The merger of accumulated rice genetic resources provides an opportunity to detect key genomic loci and candidate genes that influence the flooding tolerance of rice. RESULTS: In the present study, a whole-genome meta-analysis was conducted on 120 quantitative trait loci (QTL) obtained from 16 independent QTL studies reported from 2004 to 2023. These QTL were confined to 18 meta-QTL (MQTL), and ten MQTL were successfully validated by independent genome-wide association studies from diverse natural populations. The mean confidence interval (CI) of the identified MQTL was 3.44 times narrower than the mean CI of the initial QTL. Moreover, four core MQTL loci with genetic distance less than 2 cM were obtained. By combining differentially expressed genes (DEG) from two transcriptome datasets with 858 candidate genes identified in the core MQTL regions, we found 38 common differentially expressed candidate genes (DECGs). In silico expression analysis of these DECGs led to the identification of 21 genes with high expression in embryo and coleoptile under submerged conditions. These DECGs encode proteins with known functions involved in submergence tolerance including WRKY, F-box, zinc fingers, glycosyltransferase, protein kinase, cytochrome P450, PP2C, hypoxia-responsive family, and DUF domain. By haplotype analysis, the 21 DECGs demonstrated distinct genetic differentiation and substantial genetic distance mainly between indica and japonica subspecies. Further, the MQTL7.1 was successfully validated using flanked marker S2329 on a set of genotypes with phenotypic variation. CONCLUSION: This study provides a new perspective on understanding the genetic basis of submergence tolerance in rice. The identified MQTL and novel candidate genes lay the foundation for marker-assisted breeding/engineering of flooding-tolerant cultivars conducive to direct seeding.

Micrometeorological monitoring reveals that canopy temperature is a reliable trait for the screening of heat tolerance in rice.

Micrometeorological monitoring is not just an effective method of determining the impact of heat stress on rice, but also a reliable way of understanding how to screen for heat tolerance in rice. The aim of this study was to use micrometeorological monitoring to determine varietal differences in rice plants grown under two weather scenarios-Long-term Heat Scenario (LHS) and Normal Weather Scenario (NWS)- so as to establish reliable methods for heat tolerance screening. Experiments were conducted with two heat susceptible varieties-Mianhui 101 and IR64-and two heat tolerant varieties, Quanliangyou 681 and SDWG005. We used staggered sowing method to ensure that all varieties flower at the same time. Our results showed that heat tolerant varieties maintained lower canopy temperature compared to heat susceptible varieties, not just during the crucial flowering period of 10 am to 12 pm, but throughout the entire day and night. The higher stomatal conductance rate observed in heat tolerant varieties possibly decreased their canopy temperatures through the process of evaporative cooling during transpiration. Conversely, we found that panicle temperature cannot be used to screen for heat tolerance at night, as we observed no significant difference in the panicle temperature of heat tolerant and heat susceptible varieties at night. However, we also reported that higher panicle temperature in heat susceptible varieties decreased spikelet fertility rate, while low panicle temperature in heat tolerant varieties increased spikelet fertility rate. In conclusion, the results of this study showed that canopy temperature is probably the most reliable trait to screen for heat tolerance in rice.

A meta-quantitative trait loci analysis identified consensus genomic regions and candidate genes associated with grain yield in rice.

Improving grain yield potential in rice is an important step toward addressing global food security challenges. The meta-QTL analysis offers stable and robust QTLs irrespective of the genetic background of mapping populations and phenotype environment and effectively narrows confidence intervals (CI) for candidate gene (CG) mining and marker-assisted selection improvement. To achieve these aims, a comprehensive bibliographic search for grain yield traits (spikelet fertility, number of grains per panicle, panicles number per plant, and 1000-grain weight) QTLs was conducted, and 462 QTLs were retrieved from 47 independent QTL research published between 2002 and 2022. QTL projection was performed using a reference map with a cumulative length of 2,945.67 cM, and MQTL analysis was conducted on 313 QTLs. Consequently, a total of 62 MQTLs were identified with reduced mean CI (up to 3.40 fold) compared to the mean CI of original QTLs. However, 10 of these MQTLs harbored at least six of the initial QTLs from diverse genetic backgrounds and environments and were considered the most stable and robust MQTLs. Also, MQTLs were compared with GWAS studies and resulted in the identification of 16 common significant loci modulating the evaluated traits. Gene annotation, gene ontology (GO) enrichment, and RNA-seq analyses of chromosome regions of the stable MQTLs detected 52 potential CGs including those that have been cloned in previous studies. These genes encode proteins known to be involved in regulating grain yield including cytochrome P450, zinc fingers, MADs-box, AP2/ERF domain, F-box, ubiquitin ligase domain protein, homeobox domain, DEAD-box ATP domain, and U-box domain. This study provides the framework for molecular dissection of grain yield in rice. Moreover, the MQTLs and CGs identified could be useful for fine mapping, gene cloning, and marker-assisted selection to improve rice productivity.

Nitrogen sources affected the biosynthesis of 2-acetyl-1-pyrroline, cooked rice elongation and amylose content in rice.

Many studies have been carried out on N sources effect on fragrant rice; however, their impact on rice grain quality is largely unclear. In this study, we evaluated the effects of different types of N sources on rice growth, yield, 2-acetyl-1-pyrroline (2AP), amylose and cooked rice elongation. Two indica rice cultivars, Basmati 385 (B385), Xiangyaxiangzhan (XYXZ) and two japonica cultivars, Yunjingyou (YJY), Daohuaxiang (DHX) were grown in experimental pots with six replications under four N sources: Potassium nitrate (KNO3), ammonium bicarbonate (NH4HCO3), urea (H2NCONH2) and sodium nitrate (NaNO3) in 2019 and 2020 early seasons. Our results showed that N dynamics regulated the number of panicles, 1000-grain weight, grain yield, 2-acetyl-1-pyrroline, amylose and cooked rice elongation across all the four treatments. The NH4HCO3 treatment significantly increased the number of panicles and grain yield across the four rice varieties compared with KNO3, H2NCONH2 and NaNO3 N sources in both 2019 and 2020 early season, The KNO3 treatment significantly showed higher 1000-grain weight in B-385, YJY, XYXZ and DHX compared to other N sources. Compared with other N sources treatment, the NH4HCO3 treatments significantly increased the 2AP contents in heading stage leaves, matured leaves and grains of B-385, YJY, XYXZ and DHX respectively. Cooked rice elongation percentage also showed significant difference in all treatments studied with KNO3 recorded the highest across the four varieties. Analysis of major enzymes and compounds such as P5C, P5CS, PDH, Pyrroline, proline and Methylglyoxal showed remarkable differences in each cultivar at heading and maturity stages with higher activity in NH4HCO3 and H2NCONH2 treatments. Similarly, in all treatments, we also observed significant increase in amylose content percentage, with NH4HCO3 having greater percentage of amylose.

Cytological Observations and Bulked-Segregant Analysis Coupled Global Genome Sequencing Reveal Two Genes Associated with Pollen Fertility in Tetraploid Rice.

Neo-tetraploid rice with high fertility is a useful germplasm for polyploid rice breeding, which was developed from the crossing of different autotetraploid rice lines. However, little information is available on the molecular mechanism underlying the fertility of neo-tetraploid rice. Here, two contrasting populations of tetraploid rice, including one with high fertility (hereafter referred to as JG) and another with low fertility (hereafter referred to as JD), were generated by crossing Huaduo 3 (H3), a high fertility neo-tetraploid rice that was developed by crossing Jackson-4x with 96025-4x, and Huajingxian74-4x (T452), a low fertility autotetraploid rice parent. Cytological, global genome sequencing-based bulked-segregant (BSA-seq) and CRISPR/Cas9 technology were employed to study the genes associated with pollen fertility in neo-tetraploid rice. The embryo sacs of JG and JD lines were normal; however, pollen fertility was low in JD, which led to scarce fertilization and low seed setting. Cytological observations displayed low pollen fertility (25.1%) and approximately 31.3 and 27.2% chromosome lagging at metaphase I and II, and 28.8 and 24.8% chromosome straggling at anaphase I and II in JD, respectively. BSA-seq of F2-3 generations and RNA-seq of F4 generation detected a common fragment, i.e., 18,915,234-19,500,000, at chromosome 7, which was comprised of 78 genes associated with fertility. Among 78 genes, 9 genes had been known to be involved in meiosis and pollen development. Two mutants ny1 (LOC_Os07g32406) and ny2 (LOC_Os07g32040) were generated by CRISPR/Cas9 knockout in neo-tetraploid rice, and which exhibited low pollen fertility and abnormal chromosome behavior. Our study revealed that two unknown genes, LOC_Os07g32406 (NY1) and LOC_Os07g32040 (NY2) play an important role in pollen development of neo-tetraploid rice and provides a new perspective about the genetic mechanisms of fertility in polyploid rice.