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In flowering plants, callose (β-1,3-glucan) plays a vital role in pollen development, especially in the separation and development of microspores. However, the molecular mechanism of callose deposition during rice pollen development remains unclear. In this study, we isolated and characterized a novel rice pollen defective mutant, non-separated microspores 1 (nsm1), which produced “dyad” or “tetrad” pollen grains. Cytological analysis indicated disrupted interstitial callose deposition at the cell plate of dyads and tetrads in nsm1 pollens. This disruption caused sporopollenin to be massively deposited outside of the junction where the interstitial callose wall connected with the peripheral callose wall, or unevenly distributed on the interstitial pollen primexine at the late meiosis stage. Consequently, an excess tectum-like layer was formed outside of the junction, connecting with the tectum of two microspores during later developmental stages, which prevented the separation of microspores. Additionally, in the linkage area, the tectum of two microspores gradually fused or degenerated, resulting in a decreased contact area between microspores and the anther locule. Therefore, the defect in callose deposition resulted in unsuccessful separation of microspores, abnormal deposition of pollen exine, and also affected the accumulation of materials in microspores, resulting in pollen semi-sterility. NSM1, encoding a callose synthase located in the Golgi body, is ubiquitously expressed in anthers with its peak expression at the young microspore stage. The in vitro enzyme activity assay confirmed that NSM1 possesses callose synthase activity, and the enzyme activity in the nsm1 mutants was significantly reduced. Phylogenetic analysis indicated that NSM1 and its orthologs play a highly conserved role in callose biosynthesis among plant species. Taken together, we propose that NSM1 plays an essential role in male meiotic callose synthesis and later pollen wall development.

Wx is the key gene that controls amylose content (AC), and various alleles have been found in rice populations. Wxb is the major allele in japonica and produces moderate AC (15~18%). It was recently found that editing the promoter of Wx could produce a series of alleles that have different Wx activities. Although some studies have edited the promoter, few studies have focused on the natural variations in Wx. Here, we used the Rice3K database to investigate variations in the Wx promoter and found that the allele Wx1764178 (A/G) has a higher LD (linkage disequilibrium) with the two key SNPs (1765751, T/G; 1768006, A/C), which could produce different Wx alleles and influence AC, as reported previously. Further study showed that the Wx1764178 allele (A/G) is functional and influences the expression of Wx positively. Editing the A allele using CRISPR‒Cas9 produced 36 and 3 bp deletions and caused a decrease in the expression of Wx. The apparent amylose content (AAC) in the edited lines was decreased by 7.09% and 11.50% compared with that of the wild type, which was the japonica variety Nipponbare with Wxb and the A allele at 1764178, while a complementary line with the G allele showed a lower AAC than the A allele with no effect on other agronomic traits. The AAC of the edited lines showed a higher increase than that of the wild type (Nipponbare, Wxb) in low-nitrogen conditions relative to high-nitrogen conditions. We also developed a dCAPS marker to identify the allele and found that the G allele has widely been used (82.95%) in japonica-bred varieties from Jiangsu Province, China. Overall, we found a functional allele (Wx1764178, A/G) in the Wx promoter that could affect AAC in japonica cultivars and be developed as markers for quality improvement in rice breeding programs.

Abstract Background Breeding of conventional and hybrid rice (Oryza sativa L.) have solved hunger problems and increased farmers' income in the world. Molecular markers have been widely used in marker-assisted breeding and identification of larger numbers of different bred varieties in the past decades. The recently developed SNP markers are applied for more stable and detectable compared with other markers. But the cost of genotyping lots SNPs is high. So, it is essential to select less representative SNPs and inexpensive detecting methods to lower the cost and accelerate variety identification and breeding process. KASP (Kompetitive Allele-Specific PCR) is a flexible method to detect the SNPs, and large number of KASP markers have been widely used in variety identification and breeding. However, the ability of less KASP markers on massive variety identification and breeding remains unknown. Results Here, 48 KASP markers were selected from 378 markers to classify and analyze 518 varieties including conventional and hybrid rice. Through analyzing the population structure, the 48 markers could almost represent the 378 markers. In terms of variety identification, the 48 KASP markers had a 100% discrimination rate in 53 conventional indica varieties and 193 hybrid varieties, while they could distinguish 89.1% conventional japonica rice from different breeding institutes. Two more markers added would increase the ratio from 68.38 to 77.94%. Additionally, the 48 markers could be used for classification of subpopulations in the bred variety. Also, 8 markers had almost completely different genotypes between japonica and indica, and 3 markers were found to be very important for japonica hybrid rice. In hybrid varieties, the heterozygosity of chromosomes 3, 6 and 11 was relatively higher than others. Conclusions Our results showed that 48 KASP markers could be used to identify rice varieties, and the panel we tested could provide a database for breeders to identify new breeding lines. Also, the specific markers we found were useful for marker-assisted breeding in rice, including conventional and hybrid.

This paper is concerned with time periodic V-shaped traveling fronts in reaction-diffusion equations with ignition time-periodic nonlinearity. Our study mainly includes two parts: in the first part we establish the existence of periodic V-shaped traveling fronts by constructing a pair of proper supersolution and subsolution, and in the second part we prove the uniqueness and global asymptotic stability of V-shaped traveling fronts. [ABSTRACT FROM AUTHOR]

In plants, large numbers of R genes, which segregate as loci with alternative alleles conferring different levels of disease resistance to pathogens, have been maintained over a long period of evolution. The reason why hosts harbor susceptible alleles in view of their null contribution to resistance is unclear. In rice, a single copy gene, Pi-ta, segregates for 2 expressed clades of alleles, 1 resistant and the other susceptible. We simulated loss-of-function of the Pi-ta susceptible allele using the CRISPR/Cas9 system to detect subsequent fitness changes and obtained insights into fitness effects related to the retention of the Pi-ta susceptible allele. Our creation of an artificial knockout of the Pi-ta susceptible allele suffered fitness-related trait declines of up to 49% in terms of filled grain yield upon the loss of Pi-ta function. The Pita susceptible alleles might serve as an off-switch to downstream immune signaling, thus contributing to the fine-tuning of plant defense responses. The results demonstrated that the susceptible Pi-ta alleles should have evolved pleiotropic functions, facilitating their retention in populations. As Pi-ta is a single copy gene with no paralogs in the genome, its function cannot be compensated by an alternative gene; whereas most other R genes form gene clusters by tandem duplications, and the function could be compensated by paralogs with high sequence similarity. This attempt to evaluate the fitness effects of the R gene in crops indicates that not all disease resistance genes incur fitness costs, which also provides a plausible explanation for how host genomes can tolerate the possible genetic load associated with a vast repertoire of R genes. [ABSTRACT FROM AUTHOR]

[Objectives] The identification of salt tolerant genetic loci in rice can provide a research basis for the molecular mechanism of salt tolerance and gene resources for improving salt tolerant cultivars. [Methods Recombinant inbred lines (RILs) derived from Zhaxima, an indica landrace variety from Yunnan Province and Nanjing 46, an elite japonica variety with superior grain quality from Jiangsu Province were used. The salt tolerance at seeding stage in the RIL population was investigated as the phenotypic value. [Results] Combined with the linkage map, a total of 4 QTLs were detected: qSST-1 , qSST-3, qSST-5 and qSST-11, located in rice chromosomes 1,3,5 and 11 , respectively. All positive alleles were from the parent Nanjing 46. Three QTLs among them were not included in chromosome intervals the same as cloned rice salt tolerance genes, and thus were described as new candidate gene loci associated with seeding-stage salt tolerance. [Conclusions] This study provides important information for further exploration and utilization of new salt tolerant QTLs in rice. It is of great significance for improving the utilization of saline land in China and ensuring the stable rice production. [ABSTRACT FROM AUTHOR]

Environmental conditions are key factors in the progression of plant disease epidemics. Light affects the outbreak of plant diseases, but the underlying molecular mechanisms are not well understood. Here, we report that the light-harvesting complex II protein, LHCB5, from rice is subject to light-induced phosphorylation during infection by the rice blast fungus Magnaporthe oryzae. We demonstrate that single-nucleotide polymorphisms (SNPs) in the LHCB5 promoter control the expression of LHCB5, which in turn correlates with the phosphorylation of LHCB5. LHCB5 phosphorylation enhances broadspectrum resistance of rice to M. oryzae through the accumulation of reactive oxidative species (ROS) in the chloroplast. We also show that LHCB5 phosphorylation-induced resistance is inheritable. Our results uncover an immunity mechanism mediated by phosphorylation of light-harvesting complex II. [ABSTRACT FROM AUTHOR]

Rice blast disease is one of the most devastating diseases in rice production, which severely affects the high and stable yield of rice. The formation of appressorium plays a key role in the pathogenesis of Magnaporthe grisea in rice. It has been confirmed that a P-type ATPase(P-ATPase) is involved in the formation of appressorium. A number of small molecular substances are able to enter the pathogen from the host during the interactions between pathogens and hosts, thus resisting the infection of pathogens. In this study, a 232 bp DNA sequence with good specificity from the first exon of P-ATPase gene MgAPT2 was used as an interference fragment and was inserted into interference vector forward and reversely.The interfering vector was then transformed into rice blast-susceptible rice variety Nipponbare via Agrobacterium-mediated transformation. Identification of rice plants inoculated with M. grisea at the seedling stage and detection of the expression level of P-ATPase gene MgAPT2 showed that the expression level of MgAPT2 gene in transgenic plants was reduced and the rice blast resistance was improved. This study provided a new way for the innovation of rice germplasm resources resistant to rice blast disease. [ABSTRACT FROM AUTHOR]

The Magnaporthe oryzae avirulence gene AvrPib is required for the resistance mediated by its cognate resistance gene Pib, which has been intensively used in indica rice breeding programs in many Asian countries. However, the sequence diversity of AvrPib among geographically distinct M. oryzae populations was recently shown to be increasing. Here, we selected a field population consisting of 248 rice blast isolates collected from a disease hotspot in Philippine for the analysis of AvrPib haplotypes and their pathogenicity against Pib. We found that all of the isolates were virulent to Pib and each of them contained an insertion of Pot3 transposon in AvrPib. Moreover, Pot3 insertion was detected in different genomic positions, resulting in three different AvrPib haplotypes, designated avrPib-Hl to H3. We further conducted a genome-wide Pot2 fingerprinting analysis by repetitive element palindromic polymerase chain reaction (PCR) and identified seven different lineages out of 47 representative isolates. The isolates belonging to the same lineage often had the same AvrPib haplotype. In contrast, the isolates having the same AvrPib haplotypes did not always belong to the same lineages. Both mating types MAT1-1 and MAT1-2 were identified in the population in Bohol and the latter appeared dominant. On the host side, we found that 32 of 52 released rice varieties in the Philippines contained Pib diagnosed by PCR gene-specific primers and DNA sequencing of gene amplicons, suggesting that it was widely incorporated in different rice varieties. Our study highlights the genetic dynamics of rice blast population at both the AvrPib locus and the genome-wide levels, providing insight into the mechanisms of the mutations in AvrPib leading to the breakdown of Pi/j-mediated resistance in rice. [ABSTRACT FROM AUTHOR]