DWARF AND LESS TILLERS ON CHROMOSOME 3 Promotes Tillering in Rice by Sustaining FLORAL ORGAN NUMBER 1 Expression.

Three key factors determine yield in rice (Oryza sativa): panicle number, grain number, and grain weight. Panicle number is strongly associated with tiller number. Although many genes regulating tillering have been identified, whether Dof proteins are involved in controlling plant architecture remains unknown. The dwarf and less tillers on chromosome 3 (dlt3) rice mutant produces fewer tillers than the wild type. We cloned DLT3, which encodes a Dof protein that interacts with MONOCULM 3 (MOC3) in vivo and in vitro and recruits MOC1, forming a DLT3-MOC3-MOC1 complex. DLT3 binds to the promoter of FLORAL ORGAN NUMBER 1 (FON1) to activate its transcription and positively regulate tiller number. The overexpression of MOC1, MOC3, or FON1 in the dlt3 mutant increased tiller number. Collectively, these results suggest a model in which DLT3 regulates tiller number by maintaining the expression of MOC1, MOC3, and FON1. We discovered that DLT3 underwent directional selection in the Xian/indica and Geng/japonica populations during rice domestication. To provide genetic resources for breeding varieties with optimal panicle numbers, we performed large-scale diversity sequencing of the 1080-bp DLT3 coding region of 531 accessions from different countries and regions. Haplotype analysis showed that the superior haplotype, DLT3H1, produced the most tillers, while haplotype DLT3H6 produced the fewest tillers. Our study provides important germplasm resources for breeding super high-yielding rice varieties with combinations of superior haplotypes in different target genes, which will help overcome the challenge of food and nutritional security in the future.

Study on the strength deterioration characteristic and damage model of coal pillar dams with repeated water immersion in underground reservoirs.

The continuous operation of coal mine underground reservoirs exposes the coal pillar dams to mining disturbances and prolonged water immersion, resulting in the deterioration of coal pillars' mechanical properties and posing a serious threat to the dam stability. To this end, coal samples from the proposed pillar dam in the 5-2 coal seam of Daliuta Mine in Shendong Mining Area were selected for conducting water absorption tests and triaxial compression tests under conditions of repeated water immersion, in order to study the deterioration of the mechanical properties and acoustic emission damage characteristic of coal samples as well as the mechanism behind the deterioration of coal samples under the water-rock interaction. The results indicated that: (1) the saturated water content of coal samples exhibited a progressive increase as the water immersion times increased, but with a diminishing rate of growth. (2) As the water immersion times increased, the compressive strength, cohesive force, and internal friction angle of coal samples gradually decreased. Notably, the deterioration effect was more pronounced in compressive strength and cohesive force, while the decline in internal friction angle was relatively minor, and the total deterioration degree and the stage deterioration degree of the above three had evident cumulativity and non-uniformity. The progressive rise in water immersion times led to a gradual attenuation of the deterioration effect. Meanwhile, the confining pressure exhibited a certain inhibitory impact on the strength deterioration of coal samples. (3) Compared to the dry coal samples, the average AE count rate of coal samples subjected to a single water immersion exhibited a significant decrease, and subsequent water immersion for two, three, and four times resulted in a very minor decrease in the average AE count rate. (4) The AE cumulative ringing counts in coal samples exhibited varying degrees of reduction as water immersion times increased. Specifically, the most significant decrease in AE cumulative ringing counts occurred after the initial water immersion, followed by a gradual decrease thereafter. The energy-releasing capacity of coal samples decreased, while their plasticity exhibited a gradual increase. (5) A damage model was developed for coal samples based on the water immersion times. The model indicated that the damage to coal samples increased as the water immersion times increased, and the damage rate gradually decreased and eventually stabilized. (6) The deterioration mechanism of coals under the water-rock interaction was explained. Through repeated water immersion, the physical, chemical, and mechanical interactions between water and coal induced alterations in the internal microstructure of coal samples, resulting in the deterioration of mechanical properties such as compressive strength, cohesive force, and internal friction angle, which was a cumulative damage process from the microscopic to the macroscopic level.

Research on fire early warning index system of coal mine goaf based on multi-parameter fusion.

In order to improve the precision of goaf fire early warning outcomes, this paper obtains the temperature characteristic index of goaf fire early warning by using a coal spontaneous combustion thermogravimetric test and a coal spontaneous combustion programmed heating test. The major gas index and auxiliary gas index for early warning are derived using the integration of the Graham coefficient and grey correlation approach. The D-S evidence theory, which involves optimizing weight allocation, is utilized to integrate the early warning temperature index and various gas indexes. Based on the fusion results, a coal mine goaf fire early warning index system is developed through multi-parameter fusion. The early warning index system is then validated through a programmed heating experiment. The results show that the process of coal spontaneous combustion can be categorized into six distinct stages: latent stage, oxidation stage, critical stage, pyrolysis stage, fission stage, and combustion stage. These stages are determined by the characteristic temperatures of coal spontaneous combustion, which are 31.7 °C, 54.8 °C, 153.7 °C, 204.5 °C, and 241.6 °C. The major gas index for early warning of goaf fires can be determined by 100∆(CO)/∆O2(%). Additionally, auxiliary gas indexes such as C3H8/CH4, C3H8/C2H6, C2H4/C2H6, and C2H2 can be used for goaf fire early warning. The programmed heating experiment shows that the early warning system software is designed by the multi-parameter fusion goaf fire early warning index system is accurate and effective. The selection of the goaf fire early warning index is more rational and precise when using the multi-parameter fusion goaf fire early warning index system based on the D-S evidence theory of weight allocation. It offers robust support for enhancing the goaf fire early warning index system and predicting coal mine goaf fires.

OsCPK12 phosphorylates OsCATA and OsCATC to regulate H2O2 homeostasis and improve oxidative stress tolerance in rice.

Calcium-dependent protein kinases (CPKs), the best-characterized calcium sensors in plants, regulate many aspects of plant growth and development as well as plant adaptation to biotic and abiotic stresses. However, how CPKs regulate the antioxidant defense system remains largely unknown. We previously found that impaired function of OsCPK12 leads to oxidative stress in rice, with more H2O2, lower catalase (CAT) activity, and lower yield. Here, we explored the roles of OsCPK12 in oxidative stress tolerance in rice. Our results show that OsCPK12 interacts with and phosphorylates OsCATA and OsCATC at Ser11. Knockout of either OsCATA or OsCATC leads to an oxidative stress phenotype accompanied by higher accumulation of H2O2. Overexpression of the phosphomimetic proteins OsCATAS11D and OsCATCS11D in oscpk12-cr reduced the level of H2O2 accumulation. Moreover, OsCATAS11D and OsCATCS11D showed enhanced catalase activity in vivo and in vitro. OsCPK12-overexpressing plants exhibited higher CAT activity as well as higher tolerance to oxidative stress. Our findings demonstrate that OsCPK12 affects CAT enzyme activity by phosphorylating OsCATA and OsCATC at Ser11 to regulate H2O2 homeostasis, thereby mediating oxidative stress tolerance in rice.

Failure characteristics and conditions of rock-coal combination structure with weak layer under dynamic and static stresses.

To comprehensively understand the mechanical response of a rock-coal combination structure containing a weak layer, a series of laboratory static loading and impact loading experiments were conducted. The results showed that under static load, the sliding process of the rock coal structure was relatively slow, and fragments can be observed. Under the action of horizontal impact loading, the whole coal stratum slipped out rapidly, and the process lasted only 0.05 s. Under the horizontal and axial impact loads, the coal stratum remained stable first, and then it slipped out as a whole under the action of static load. Additionally, a sliding instability criterion of a rock coal structure containing a weak layer was established based on theoretical analysis. The key parameter P value was checked through a numerical simulation experiment. It was found that the value was linearly related to the mechanical properties of the weak layer and overburden stress, and the experimental results coincided with the theoretical results. Finally, the relationship between sliding rockburst and strain rockburst was discussed, and these results can provide an important scientific basis for the prevention and control of dynamic disasters in deep mining.

Research on the dynamic evolution law of fissures in shallow-buried and short-distance coal seam mining in Lijiahao Coal Mine.

Aiming at the problem of spontaneous combustion of coal relics caused by the overburden fracture network penetrating the upper and lower coal seams in the process of shallow-buried and short-distance coal seam mining, the 31114 working face of Lijiahao coal mine was used as the research background to study the characteristics of overburden transport and fracture development in shallow-buried and short-distance coal seam mining by using physical similar simulation test; the fractal dimension and image processing techniques were used to quantify the overburden fractures; the classical mechanical models of "solid support beam" and "masonry beam" were combined to analyze the causes of fracture dynamic evolution. The results show that: (1) Before the key seam fracture, the stress in the upper rock seam only changes in a small amount, and the stress in the lower rock seam evolves similarly to the single coal seam mining; when the key seam fracture is broken, the stress in the upper and lower rock seams will change by jumps. (2) The fractal dimension of the fissures rised from 1.4 to 1.5, the total area of fissures is increased from 16,638 pixels to 17,707 pixels, and the total length is increased from 2217 to 3071 pixels; after the main key layer of the overlying rock is broken, the fractal dimension of the fissures is reduced from 1.56 to 1.5, and the total area of fissures is reduced from 31,451 pixels to 29,089 pixels, the total length has increased from 5657 to 6619 pixels. (3) Before the key layer between the coal seams is broken, it will be suspended to form a "fixed beam". After the first break, the broken rock above it will settle synchronously until the rock blocks form a hinged structure and then collapse. After the fall stops, the key layer periodically breaks to form a "masonry beam" structure, and the overlying stratum settles synchronously.

Mutation of DEFECTIVE EMBRYO SAC1 results in a low seed-setting rate in rice by regulating embryo sac development.

The seed-setting rate has a significant effect on grain yield in rice (Oryza sativa L.). Embryo sac development is essential for seed setting; however, the molecular mechanism underlying this process remains unclear. Here, we isolated defective embryo sac1 (des1), a rice mutant with a low seed-setting rate. Cytological examination showed degenerated embryo sacs and reduced fertilization capacity in des1. Map-based cloning revealed a nonsense mutation in OsDES1, a gene that encodes a putative nuclear envelope membrane protein (NEMP)-domain-containing protein that is preferentially expressed in pistils. The OsDES1 mutation disrupts the normal formation of functional megaspores, which ultimately results in a degenerated embryo sac in des1. Reciprocal crosses showed that fertilization is abnormal and that the female reproductive organ is defective in des1. OsDES1 interacts with LONELY GUY (LOG), a cytokinin-activating enzyme that acts in the final step of cytokinin synthesis; mutation of LOG led to defective female reproductive organ development. These results demonstrate that OsDES1 functions in determining the rice seed-setting rate by regulating embryo sac development and fertilization. Our study sheds light on the function of NEMP-type proteins in rice reproductive development.

qHD5 encodes an AP2 factor that suppresses rice heading by down-regulating Ehd2 expression.

Heading date is crucial for rice reproduction and the geographical expansion of cultivation. We fine-mapped qHD5 and identified LOC_Os05g03040, a gene that encodes an AP2 transcription factor, as the candidate gene of qHD5 in our previous study. In this article, using two near-isogenic lines NIL(BG1) and NIL(XLJ), which were derived from the progeny of the cross between BigGrain1 (BG1) and Xiaolijing (XLJ), we verified that LOC_Os05g03040 represses heading date in rice through genetic complementation and CRISPR/Cas9 gene-editing experiments. Complementary results showed that qHD5 is a semi-dominant gene and that the qHD5XLJ and qHD5BG1 alleles are both functional. The homozygous mutant line generated from knocking out qHD5XLJ in NIL(XLJ) headed earlier than NIL(XLJ) under both short-day and long-day conditions. In addition, the homozygous mutant line of qHD5BG1 in NIL(BG1) also headed slightly earlier than NIL(BG1). All of these results show that qHD5 represses the heading date in rice. Transient expression showed that the qHD5 protein localizes to the nucleus. Transactivation activity assays showed that the C-terminus is the critical site that affects self-activation in qHD5XLJ. qRT-PCR analysis revealed that qHD5 represses flowering by down-regulating Ehd2. qHD5 may have been selected during indica rice domestication.

Disruption of OsPHD1, Encoding a UDP-Glucose Epimerase, Causes JA Accumulation and Enhanced Bacterial Blight Resistance in Rice.

Lesion mimic mutants (LMMs) have been widely used in experiments in recent years for studying plant physiological mechanisms underlying programmed cell death (PCD) and defense responses. Here, we identified a lesion mimic mutant, lm212-1, which cloned the causal gene by a map-based cloning strategy, and verified this by complementation. The causal gene, OsPHD1, encodes a UDP-glucose epimerase (UGE), and the OsPHD1 was located in the chloroplast. OsPHD1 was constitutively expressed in all organs, with higher expression in leaves and other green tissues. lm212-1 exhibited decreased chlorophyll content, and the chloroplast structure was destroyed. Histochemistry results indicated that H2O2 is highly accumulated and cell death is occurred around the lesions in lm212-1. Compared to the wild type, expression levels of defense-related genes were up-regulated, and resistance to bacterial pathogens Xanthomonas oryzae pv. oryzae (Xoo) was enhanced, indicating that the defense response was activated in lm212-1, ROS production was induced by flg22, and chitin treatment also showed the same result. Jasmonic acid (JA) and methyl jasmonate (MeJA) increased, and the JA signaling pathways appeared to be disordered in lm212-1. Additionally, the overexpression lines showed the same phenotype as the wild type. Overall, our findings demonstrate that OsPHD1 is involved in the regulation of PCD and defense response in rice.

Dissection of Closely Linked Quantitative Trait Locis Controlling Grain Size in Rice.

Grain size is a key constituent of grain weight and appearance in rice. However, insufficient attention has been paid to the small-effect quantitative trait loci (QTLs) on the grain size. In the present study, residual heterozygous populations were developed for mapping two genetically linked small-effect QTLs for grain size. After the genotyping and the phenotyping of five successive generations, qGS7.1 was dissected into three QTLs and two were selected for further analysis. The qTGW7.2a was finally mapped into a 21.10 kb interval containing four annotated candidate genes. Transcript levels assay showed that the expression of the candidates LOC_Os07g39490 and the LOC_Os07g39500 were significantly reduced in the NIL-qTGW7.2aBG1 . The cytological observation indicated that qTGW7.2a regulated the grain width through controlling the cell expansion. Using the same strategy, qTGW7.2b was fine-mapped into a 52.71 kb interval containing eight annotated candidate genes, showing a significant effect on the grain length and width with opposite allelic directions, but little on the grain weight. Our study provides new genetic resources for yield improvement and for fine-tuning of grain size in rice.

Rice dwarf and low tillering 10 (OsDLT10) regulates tiller number by monitoring auxin homeostasis.

Tiller number is a crucial agronomic trait that directly affects the number of effective panicles and yield formation in rice. Here, we report a semi-dwarf and low tillering mutant Osdlt10 (dwarf and low tillering 10) that exhibited reduced tiller number, semi-dwarfism, increased grain width, low seed-setting rate, curled leaf tip and a series of abnormalities of agronomic traits. Phenotypic observations showed that Osdlt10 mutants had defects in tiller bud formation and grew slowly at the tillering stage. Map-based cloning revealed that LOC_Os10g41310 was the responsible gene for OsDLT10, which was subsequently demonstrated using the CRISPR/Cas9 system and a complementary experiment. Expression pattern analysis indicated that OsDLT10 was primarily expressed in the stem node, the basic part of axillary bud and leaf sheath, pulvinus. The hormone treatment investigation indicated that extremely high of exogenous auxin concentrations can inhibit the expression of OsDLT10. Endogenous auxin content decreased significantly at the base of stem node and axillary bud in Osdlt10 mutants. The results showed that OsDLT10 was related to auxin. qPCR analysis results further showed that the expression levels of auxin transport genes (PINs) and early response genes (IAAs) were significantly increased. The expression levels of WUS-like and FON1 were substantially decreased in the Osdlt10 mutants. These results revealed that OsDLT10 played a critical role in influencing tiller number, likely in association with hormone signals and the WUS-CLV pathway, to regulate axillary bud development in rice.

ES5 is involved in the regulation of phosphatidylserine synthesis and impacts on early senescence in rice (Oryza sativa L.).

Leaf senescence, which affects plant growth and yield in rice, is an ideal target for crop improvement and remarkable advances have been made to identify the mechanism underlying this process. We have characterized an early senile mutant es5 (early leaf senescence 5) in rice exhibiting leaf yellowing phenotype after the 4-leaf stage. This phenotype was confirmed by the higher accumulation of reactive oxygen species (ROS) and malondialdehyde (MDA), the disintegration of chloroplasts, reduction in chlorophyll content and photosynthetic rate and up-regulation of senescence-associated genes (SAGs) like Osh36, OsI57, and OsI85. Positional cloning revealed that the es5 phenotype is the result of one base substitution in ES5, encoding phosphatidylserine synthase (PSS) family protein, which is involved in the base-exchange type reaction to synthesize the minor membrane phospholipid phosphatidylserine. Functional complementation of ES5 in the es5 plants completely restored the wild-type phenotype. Ultra-high-performance liquid chromatography (UHPLC) analysis showed that es5 plants had increased levels of phosphatidylserine (PS) and decreased level of phosphatidylcholine (PC). These results provide evidence about the role of PS in rice leaf senescence.

Impaired Function of the Calcium-Dependent Protein Kinase, OsCPK12, Leads to Early Senescence in Rice (Oryza sativa L.).

Premature leaf senescence affects plant yield and quality, and numerous researches about it have been conducted until now. In this study, we identified an early senescent mutant es4 in rice (Oryza sativa L.); early senescence appeared approximately at 60 dps and became increasingly senescent with the growth of es4 mutant. We detected that content of reactive oxygen species (ROS) and malondialdehyde (MDA), as well as activity of superoxide dismutase (SOD) were elevated, while chlorophyll content, soluble protein content, activity of catalase (CAT), activity of peroxidase (POD) and photosynthetic rate were reduced in the es4 mutant leaves. We mapped es4 in a 33.5 Kb physical distance on chromosome 4 by map-based cloning. Sequencing analysis in target interval indicated there was an eight bases deletion mutation in OsCPK12 which encoded a calcium-dependent protein kinase. Functional complementation of OsCPK12 in es4 completely restored the normal phenotype. We used CRISPR/Cas9 for targeted disruption of OsCPK12 in ZH8015 and all the mutants exhibited the premature senescence. All the results indicated that the phenotype of es4 was caused by the mutation of OsCPK12. Overexpression of OsCPK12 in ZH8015 enhanced the net photosynthetic rate (P n) and chlorophyll content. OsCPK12 was mainly expressed in green organs. The results of qRT-PCR analysis showed that the expression levels of some key genes involved in senescence, chlorophyll biosynthesis, and photosynthesis were significantly altered in the es4 mutant. Our results demonstrate that the mutant of OsCPK12 triggers the premature leaf senescence; however, the overexpression of OsCPK12 may delay its growth period and provide the potentially positive effect on productivity in rice.

Genetic Dissection of qPCG1 for a Quantitative Trait Locus for Percentage of Chalky Grain in Rice (Oryza sativa L.).

Rice is a pivotal cereal crop that provides the staple food for more than half of the world's population. Along with improvements in the standard of living, people not only pay attention to the grain yield but also to the grain quality. Chalkiness is one of the most important index of grain quality. In this study, qPCG1, a QTL for percentage of chalky grain, was mapped in an interval with a physical distance about 139 kb on chromosome 1 by residual heterozygous line (RHL) method. qPCG1 was incomplete dominant and the additive effect plays a major role and explained 6.8-21.9% of phenotypic variance within the heterogeneous region on chromosome 1. The effect of allele from Zhonghui9308 was decreasing the percentage of chalky grains (PCG). Microscope observation results indicated that there are great differences in the shape, structure and arrangement of starch granule between the chalky part and transparent part. Analysis of starch physicochemical properties showed that the total starch content, amylose content and chain length distribution of amylopectin changed while the protein contents were not apparently affected with the changed chalkiness. qPCG1 had little influence on main agronomic traits and it might be useful in rice breeding for it did not bring negative effect on grain yield while reducing the chalkiness.