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Catalase negatively regulates plant immunity and is targeted and degraded by ubiquitin E3 ligase.

Phenolamides (PAs), a diverse group of specialized metabolites, including hydroxycinnamoylputrescine (HP), hydroxycinnamoylagmatine, and hydroxycinnamoyltryptamine, are important in plant resistance to biotic stress. However, the genes involved in the biosynthesis and modulation of PAs have not been fully elucidated. This study identified an HP biosynthetic gene cluster in rice (Oryza sativa) comprising one gene (OsODC) encoding a decarboxylase and two tandem-duplicated genes (OsPHT3 and OsPHT4) encoding putrescine hydroxycinnamoyl acyltransferases coexpressed in different tissues. OsODC catalyzes the conversion of ornithine to putrescine, which is used in HP biosynthesis involving OsPHT3 and OsPHT4. OsPHT3 or OsPHT4 overexpression causes HP accumulation and cell death and putrescine hydroxycinnamoyl acyltransferases (PHT) activity-dependent resistance against the fungal pathogen Magnaporthe oryzae. OsODC overexpression plants also confer enhanced resistance to M. oryzae. Notably, the basic leucine zipper transcription factor APIP5, a negative regulator of cell death, directly binds to the OsPHT4 promoter, repressing its transcription. Moreover, APIP5 suppression induces OsPHT4 expression and HP accumulation. Comparative genomic analysis revealed that the HP biosynthetic gene cluster is conserved in monocots. These results characterized a previously unidentified monocot-specific gene cluster that is involved in HP biosynthesis and contributes to defense and cell death in rice.

BackgroundBalancing the yield, quality and resistance to disease is a daunting challenge in crop breeding due to the negative relationship among these traits. Large-scale genomic landscape analysis of germplasm resources is considered to be an efficient approach to dissect the genetic basis of the complex traits. Central China is one of the main regions where thejaponicarice is produced. However, dozens of high-yield rice varieties in this region still exist with low quality or susceptibility to blast disease, severely limiting their application in rice production.ResultsHere, we re-sequence 200japonicarice varieties grown in central China over the past 30 years and analyze the genetic structure of these cultivars using 2.4 million polymorphic SNP markers. Genome-wide association mapping and selection scans indicate that strong selection for high-yield and taste quality associated with low-amylose content may have led to the loss of resistance to the rice blast fungusMagnaporthe oryzae. By extensive bioinformatic analyses of yield components, resistance to rice blast, and taste quality, we identify several superior alleles for these traits in the population. Based on this information, we successfully introduce excellent taste quality and blast-resistant alleles into the background of two high-yield cultivars and develop two elite lines, XY99 and JXY1, with excellent taste, high yield, and broad-spectrum of blast resistance.ConclusionsThis is the first large-scale genomic landscape analysis ofjaponicarice varieties grown in central China and we demonstrate a balancing of multiple agronomic traits by genomic-based strategy.

Dysfunction of susceptibility genes leads to broad-spectrum resistance (BSR) in plants. A rice ubiquitin E3 ligase degrades an ortholog of plant thermosensor to positively regulate blast resistance. Gao et al. recently showed that this E3 ligase also ubiquitinates a Ca

Plant metabolites are critical components of immune signaling pathways; however, how these small molecules contribute to plant immunity remains largely elusive. Emerging evidence demonstrates that the rice nucleotide-binding leucine-rich repeat receptor (NLR)-interacting proteins regulate the biosynthesis of ethylene, hydroxycinnamoylputrescines and diterpenoid phytoalexins to modulate plant immunity.

Nucleotide-binding leucine-rich repeat (NLR) proteins play critical roles in plant immunity. However, how NLRs are regulated and activate defense signaling is not fully understood. The rice (Oryza sativa) NLR receptor Piz-t confers broad-spectrum resistance to the fungal pathogen Magnaporthe oryzae and the RING-type E3 ligase AVRPIZ-T INTERACTING PROTEIN 10 (APIP10) negatively regulates Piz-t accumulation. In this study, we found that APIP10 interacts with two rice transcription factors, VASCULAR PLANT ONE-ZINC FINGER 1 (OsVOZ1) and OsVOZ2, and promotes their degradation through the 26S proteasome pathway. OsVOZ1 displays transcriptional repression activity while OsVOZ2 confers transcriptional activation activity in planta. The osvoz1 and osvoz2 single mutants display modest but opposite M. oryzae resistance in the non-Piz-t background. However, the osvoz1 osvoz2 double mutant exhibits strong dwarfism and cell death, and silencing of both genes via RNA interference also leads to dwarfism, mild cell death, and enhanced resistance to M. oryzae in the non-Piz-t background. Both OsVOZ1 and OsVOZ2 interact with Piz-t. Double silencing of OsVOZ1 and OsVOZ2 in the Piz-t background decreases Piz-t protein accumulation and transcription, reactive oxygen species-dependent cell death, and resistance to M. oryzae containing AvrPiz-t. Taken together, these results indicate that OsVOZ1 and OsVOZ2 negatively regulate basal defense but contribute positively to Piz-t-mediated immunity.

Plant diseases reduce crop yields and threaten global food security, making the selection of disease-resistant cultivars a major goal of crop breeding. Broad-spectrum resistance (BSR) is a desirable trait because it confers resistance against more than one pathogen species or against the majority of races or strains of the same pathogen. Many BSR genes have been cloned in plants and have been found to encode pattern recognition receptors, nucleotide-binding and leucine-rich repeat receptors, and defense-signaling and pathogenesis-related proteins. In addition, the BSR genes that underlie quantitative trait loci, loss of susceptibility and nonhost resistance have been characterized. Here, we comprehensively review the advances made in the identification and characterization of BSR genes in various species and examine their application in crop breeding. We also discuss the challenges and their solutions for the use of BSR genes in the breeding of disease-resistant crops.

Many transcription factors (TFs) in animals bind to both DNA and mRNA, regulating transcription and mRNA turnover. However, whether plant TFs function at both the transcriptional and post-transcriptional levels remains unknown. The rice (Oryza sativa) bZIP TF AVRPIZ-T-INTERACTING PROTEIN 5 (APIP5) negatively regulates programmed cell death and blast resistance and is targeted by the effector AvrPiz-t of the blast fungus Magnaporthe oryzae. We demonstrate that the nuclear localization signal of APIP5 is essential for APIP5-mediated suppression of cell death and blast resistance. APIP5 directly targets two genes that positively regulate blast resistance: the cell wall-associated kinase gene OsWAK5 and the cytochrome P450 gene CYP72A1. APIP5 inhibits OsWAK5 expression and thus limits lignin accumulation; moreover, APIP5 inhibits CYP72A1 expression and thus limits reactive oxygen species production and defense compounds accumulation. Remarkably, APIP5 acts as an RNA-binding protein to regulate mRNA turnover of the cell death- and defense-related genes OsLSD1 and OsRac1. Therefore, APIP5 plays dual roles, acting as TF to regulate gene expression in the nucleus and as an RNA-binding protein to regulate mRNA turnover in the cytoplasm, a previously unidentified regulatory mechanism of plant TFs at the transcriptional and post-transcriptional levels.

Background Rice blast is one of the most widespread and devastating diseases in rice production. Tremendous success has been achieved in identification and characterization of genes and quantitative trait loci (QTLs) conferring seedling blast resistance, however, genetic studies on panicle blast resistance have lagged far behind. Results In this study, two advanced backcross inbred sister lines (MSJ13 and MSJ18) were obtained in the process of introducing Pigm into C134S, and showed significant differences in the panicle blast resistance. One F2 population derived from the crossing MSJ13/MSJ18 was used to QTL mapping for panicle blast resistance using Genotyping by Sequencing (GBS) method. A total of 7 QTLs were identified, including a major QTL qPBR10-1 on chromosome 10 that explaining 24.21% of phenotypic variance with LOD scores of 6.62. Furthermore, qPBR10-1 was verified via the BC1F2 and BC1F3 population and narrowed to a 60.6-kb region with six candidate genes predicted, including two genes encoding exonuclease family protein, two genes encoding hypothetical protein, and two genes encoding transposon protein. The nucleotide variations and the expression patterns of the candidate genes were identified and analyzed between MSJ13 and MSJ18 through sequence comparison and RT-PCR approach, and results indicated that ORF1 and ORF2 encoding exonuclease family protein might be the causal candidate genes for panicle blast resistance in the qPBR10-1 locus. Conclusions A total of 7 QTLs conferring panicle blast resistance was identified from one F2 population derived from the crossing between two advanced backcross inbred sister lines MSJ13 and MSJ18, which harbored the broad-spectrum resistance gene Pigm. A major QTL qPBR10-1 was fine mapped in a 60.6-kb region with six candidate genes predicted, and ORF1 and ORF2 encoding exonuclease family protein might be the causal candidate genes for panicle blast resistance in the qPBR10-1 locus through sequence comparison and RT-PCR approach.

Phenolamide (PA) metabolites play important roles in the interaction between plants and pathogens. The putrescine hydroxycinnamoyl transferase genes OsPHT3 and OsPHT4 positively regulate rice cell death and resistance to Magnaporthe oryzae. The bZIP transcription factor APIP5, a negative regulator of cell death and rice immunity, directly binds to the OsPHT4 promoter to regulate putrescine-derived PAs. Whether other hydroxycinnamoyl transferase (HT) genes also participate in APIP5-mediated immunity remains unclear. Surprisingly, we find that genes encoding agmatine hydroxycinnamoyl transferases OsAHT1 and OsAHT2, tryptamine hydroxycinnamoyl transferases OsTBT1 and OsTBT2, and tyramine hydroxycinnamoyl transferases OsTHT1 and OsTHT2, responsible for the biosynthesis of polyamine-derived PAs are all up-regulated in APIP5-RNAi transgenic plants compared with segregated wild-type rice. Furthermore, both OsAHT1/2 and OsTBT1/2 are induced during M. oryzae infection, showing expression patterns similar to those previously reported for OsTHT1/2 and OsPHT3/4. Transgenic plants overexpressing either OsAHT2-GFP or OsTBT1-GFP show enhanced resistance against M. oryzae and accumulated more PA metabolites and lignin compared with wild-type plants. Interestingly, as demonstrated for OsPHT4, APIP5 directly binds to the promoters of OsAHT1/2, OsTBT1/2, and OsTHT1/2, repressing their transcription. Together, these results indicate that the HT genes are common targets of APIP5 and that PAs play critical roles in rice immunity.

Mitochondria are energy factories of cells and are important for intracellular interactions with other organelles. Emerging evidence indicates that mitochondria play essential roles in the response to pathogen infection. During infection, pathogens deliver numerous enzymes and effectors into host cells, and some of these effectors target mitochondria, altering mitochondrial morphology, metabolism, and functions. To defend against pathogen attack, mitochondria are actively involved in changing intracellular metabolism, hormone-mediated signaling, and signal transduction, producing reactive oxygen species and reactive nitrogen species and triggering programmed cell death. Additionally, mitochondria coordinate with other organelles to integrate and amplify diverse immune signals. In this review, we summarize recent advances in understanding how mitochondria function in plant immunity and how pathogens target mitochondria for host defense suppression.

Activation of plant respiratory burst oxidase homologs (RBOHs) to generate reactive oxygen species (ROS) is a crucial defense signaling event. RBOH activation occurs predominantly through N-terminal phosphorylation and the binding of a small GTPase. Two recent papers reported that C-terminal phosphorylation and ubiquitination modulates AtRBOHD activity, which extends our understanding of the fine-tuning of RBOH signaling in plant immunity.

Plant diseases severely reduce crop yields and threaten global food security. Broad-spectrum resistance (BSR) is a desirable trait because it confers resistance against more than one pathogen species or the majority of races/strains of the same pathogen. To control plant diseases, breeders have selected BSR to reduce disease occurrence and prolong the life-span of newly released cultivars in the last several decades (Mundt, Phytopathology 108(7):792–802, 2018). Although effective, breeding of BSR cultivars in crop plants is still time-consuming and technically challenging. Recently, new gene-editing technologies such as CRISPR/Cas9 have dramatically accelerated the process of plant breeding and provided an approach for rapidly creating new varieties with BSR and other beneficial traits (Borrelli et al., Front Plant Sci 9:1245, 2018). In addition, close surveillance of pathogen populations in the field can provide useful information for the deployment of appropriate resistance genes in the target regions. In this mini-review, we focus on the significance and application of the exciting results from two recent companion papers published in Nature Biotechnology that provide new strategies to develop crop plants with BSR against pathogens through targeted promoter editing of susceptibility genes in plants as well as pathogen population monitoring.

Rice blast and bacterial blight, caused by the fungus Magnaporthe oryzae and the bacterium Xanthomonas oryzae pv. oryzae (Xoo), respectively, are devastating diseases affecting rice. Here, we report that a rice valine-glutamine (VQ) motif-containing protein, OsVQ25, balances broad-spectrum disease resistance and plant growth by interacting with a U-Box E3 ligase, OsPUB73, and a transcription factor, OsWRKY53. We show that OsPUB73 positively regulates rice resistance against M. oryzae and Xoo by interacting with and promoting OsVQ25 degradation via the 26S proteasome pathway. Knockout mutants of OsVQ25 exhibit enhanced resistance to both pathogens without a growth penalty. Furthermore, OsVQ25 interacts with and suppresses the transcriptional activity of OsWRKY53, a positive regulator of plant immunity. OsWRKY53 downstream defense-related genes and brassinosteroid signaling genes are upregulated in osvq25 mutants. Our findings reveal a ubiquitin E3 ligase-VQ protein-transcription factor module that fine-tunes plant immunity and growth at the transcriptional and posttranslational levels.

Additional file of Deep sequencing reveals the complex and coordinated transcriptional regulation of genes related to grain quality in rice cultivars

A fungal effector targets a heat shock–dynamin protein complex to modulate mitochondrial dynamics and reduce plant immunity., Mitochondria are essential for animal and plant immunity. Here, we report that the effector MoCDIP4 of the fungal pathogen Magnaporthe oryzae targets the mitochondria-associated OsDjA9-OsDRP1E protein complex to reduce rice immunity. The DnaJ protein OsDjA9 interacts with the dynamin-related protein OsDRP1E and promotes the degradation of OsDRP1E, which functions in mitochondrial fission. By contrast, MoCDIP4 binds OsDjA9 to compete with OsDRP1E, resulting in OsDRP1E accumulation. Knockout of OsDjA9 or overexpression of OsDRP1E or MoCDIP4 in transgenic rice results in shortened mitochondria and enhanced susceptibility to M. oryzae. Overexpression of OsDjA9 or knockout of OsDRP1E in transgenic rice, in contrast, leads to elongated mitochondria and enhanced resistance to M. oryzae. Our study therefore reveals a previously unidentified pathogen-infection strategy in which the pathogen delivers an effector into plant cells to target an HSP40-DRP complex; the targeting leads to the perturbation of mitochondrial dynamics, thereby inhibiting mitochondria-mediated plant immunity.

Crop diseases are major factors responsible for substantial yield losses worldwide, which affects global food security. The use of resistance (R) genes is an effective and sustainable approach to controlling crop diseases. Here, we review recent advances on R gene studies in the major crops and related wild species. Current understanding of the molecular mechanisms underlying R gene activation and signaling, and susceptibility (S) gene-mediated resistance in crops are summarized and discussed. Furthermore, we propose some new strategies for R gene discovery, how to balance resistance and yield, and how to generate crops with broad-spectrum disease resistance. With the rapid development of new genome-editing technologies and the availability of increasing crop genome sequences, the goal of breeding next-generation crops with durable resistance to pathogens is achievable, and will be a key step toward increasing crop production in a sustainable way.

Phenylalanine ammonia lyases (PALs) are involved in resistance to plant pathogens, but the mechanism by which they contribute to plant immunity is unclear. Two recent studies provide direct evidence for PALs' contributions to rice broad-spectrum resistance (BSR) mediated by nucleotide-binding domain and leucine-rich repeat-containing (NLR) proteins and RNA-binding proteins (Zhai et al., Mol. Cell, 2019; Zhou et al., Proc. Natl. Acad. Sci. U. S. A., 2018).

Crop diseases cause enormous yield losses and threaten global food[ED1] security. The use of highly resistant cultivars can effectively control plant diseases, but in crops, genetic immunity to disease often comes with an unintended reduction in growth and yield. Here, we review recent advances in understanding how nucleotide-binding domain, leucine-rich repeat (NLR) receptors and cell wall-associated kinase (WAK) proteins function in balancing immunity and yield. We also discuss the role of plant hormones and transcription factors in regulating the trade-offs between plant growth and immunity. Finally, we describe how a novel mechanism of translational control of defense proteins can enhance immunity without the reduction in fitness.

Additional file 1:Filter results of the five MPSS and SBS libraries. A) A total of 39,288 distinct 17-base expressed signatures from the five MPSS libraries were processed according to three filters: significance, reliability, and genomic match. B) Similarly, 397,543 signatures from the five SBS libraries were also processed using these same filters as previously described by Meyers et al. [45]. (PPT 134 KB)

Additional file 2:Classification of the MPSS and SBS signatures from the five libraries based on their location on the annotated gene (hits = 1) (See Meyers et al. 2004[45]for details).(DOC 60 KB)

Authors’ original file for figure 2

Additional file 7:Conserved cis elements in the promoter region of the highly induced genes (≥50 fold) in Cypress (compared to LaGrue and Nipponbare) and Ilpumbyeo (compared to YR15965 and Nipponbare) that are involved in seed development. (DOC 46 KB)

Authors’ original file for figure 1

A central goal of crop improvement is to breed varieties with broad-spectrum resistance (BSR) to pathogens, but most of the major resistance (R) genes identified to date confer race-specific resistance to their adapted pathogens. Although these R genes are effective for a specific pathogen, their durability in the field is typically short due to mutations in the pathogen population that overcome the resistance. An alternative strategy of incorporating multiple R genes against different pathogens into elite cultivars is time-consuming and technically challenging, and usually results in a yield penalty as the crop diverts energy to implementing disease resistance (1). Many pathogens infect rice ( Oryza sativa ), the staple food crop of over half of the world’s population. Among them, the fungal pathogen Magnaporthe oryzae and the bacterial pathogen Xanthomonas oryzae pv oryzae ( Xoo ) are two of the most destructive and can cause devastating yield losses in most rice-growing countries (2). Over the past two decades, many rice R genes have been identified, but none confers resistance to both pathogens. Although manipulating the expression of several defense-responsive genes, or genes in defense signaling pathways, has led to BSR (3, 4), few such genes have been successfully deployed in rice production for disease control. In PNAS, Zhou et al. (5) report the identification of the broad-spectrum resistance Kitaake-1 ( Bsr-k1 ) gene, which negatively regulates BSR, and the bsr-k1 allele, which confers nonspecific BSR to both M. oryzae and Xoo without a yield penalty. The resistant bsr-k1 mutant was identified from an ethylmethane sulfonate-treated mutant population of the japonica cultivar Kitaake that had been inoculated with seven Kitaake-compatible M. oryzae … [↵][1]1To whom correspondence should be addressed. Email: wang.620{at}osu.edu. [1]: #xref-corresp-1-1

Summary Bowman–Birk trypsin inhibitors (BBIs) play important roles in animal and plant immunity, but how these protease inhibitors are involved in the immune system remains unclear. Here, we show that the rice (Oryza sativa) BBI protein APIP4 is a common target of a fungal effector and an NLR receptor for innate immunity. APIP4 exhibited trypsin inhibitor activity in vitro and in vivo. Knockout of APIP4 in rice enhanced susceptibility, and overexpression of APIP4 increased resistance to the fungal pathogen Magnaporthe oryzae. The M. oryzae effector AvrPiz‐t interacted with APIP4 and suppressed APIP4 trypsin inhibitor activity. By contrast, the rice NLR protein Piz‐t interacted with APIP4, enhancing APIP4 transcript and protein levels, and protease inhibitor activity. Our findings reveal a novel host defence mechanism in which a host protease inhibitor targeted by a fungal pathogen is protected by an NLR receptor.

Summary Because of the frequent breakdown of major resistance (R) genes, identification of new partial R genes against rice blast disease is an important goal of rice breeding. In this study, we used a core collection of the Rice Diversity Panel II (C‐RDP‐II), which contains 584 rice accessions and are genotyped with 700 000 single‐nucleotide polymorphism (SNP) markers. The C‐RDP‐II accessions were inoculated with three blast strains collected from different rice‐growing regions in China. Genome‐wide association study identified 27 loci associated with rice blast resistance (LABRs). Among them, 22 LABRs were not associated with any known blast R genes or QTLs. Interestingly, a nucleotide‐binding site leucine‐rich repeat (NLR) gene cluster exists in the LABR12 region on chromosome 4. One of the NLR genes is highly conserved in multiple partially resistant rice cultivars, and its expression is significantly up‐regulated at the early stages of rice blast infection. Knockout of this gene via CRISPR‐Cas9 in transgenic plants partially reduced blast resistance to four blast strains. The identification of this new non‐strain specific partial R gene, tentatively named rice blast Partial Resistance gene 1 (PiPR1), provides genetic material that will be useful for understanding the partial resistance mechanism and for breeding durably resistant cultivars against blast disease of rice.

Lesion mimic mutants are ideal genetic materials to study programmed cell death and defense signaling in plants. However, the molecular basis of lesion mimic formation remains largely unknown. Here, we first used a proteomic approach to identify differentially expressed proteins during dynamic lesion mimic formation in the rice oscul3a mutant, then electron microscope observation and physiological assays were used to analyze the mutant. The oscul3a mutant had disrupted cell metabolism balance, and the identified differentially expressed proteins were mainly located in the chloroplast and cytoplasm, which caused enhanced lipid metabolism, but suppressed carbon/nitrogen metabolism with reduced growth and grain quality. The oscul3a mutant had higher salicylic acid (SA) concentration in leaves, and H2O2 was shown to accumulate late in the formation of lesions. The secondary metabolite coumarin induced reactive oxygen species (ROS) and had rice blast resistance activity. Moreover, the cell death initiated lesion mimic formation of oscul3a mutant was light-sensitive, which might be associated with metabolite biosynthesis and accumulation. This study sheds light on the metabolic transition associated with cell death and defense response, which is under tight regulation by OsCUL3a and metabolism-related proteins, and the newly identified chemicals in the secondary metabolic pathway can potentially be used to control disease in crop plants.

Cullin3-based RING E3 ubiquitin ligases (CRL3), composed of Cullin3 (CUL3), RBX1, and BTB proteins, are involved in plant immunity, but the function of CUL3 in the process is largely unknown. Here, we show that rice (Oryza sativa) OsCUL3a is important for the regulation of cell death and immunity. The rice lesion mimic mutant oscul3a displays a significant increase in the accumulation of flg22- and chitin-induced reactive oxygen species, and in pathogenesis-related gene expression as well as resistance to Magnaporthe oryzae and Xanthomonas oryzae pv oryzae. We cloned the OsCUL3a gene via a map-based strategy and found that the lesion mimic phenotype of oscul3a is associated with the early termination of OsCUL3a protein. Interaction assays showed that OsCUL3a interacts with both OsRBX1a and OsRBX1b to form a multisubunit CRL in rice. Strikingly, OsCUL3a interacts with and degrades OsNPR1, which acts as a positive regulator of cell death in rice. Accumulation of OsNPR1 protein is greater in the oscul3a mutant than in the wild type. Furthermore, the oscul3a osnpr1 double mutant does not exhibit the lesion mimic phenotype of the oscul3a mutant. Our data demonstrate that OsCUL3a negatively regulates cell death and immunity by degrading OsNPR1 in rice.

Rice (Oryza sativa) is not only the staple food for half of the world's population but also a model monocot plant for molecular biology studies. Although rice genes have been extensively investigated in the last two decades, the functions of many genes in the rice genome are still not known. One of the rapid and efficient approaches for determining gene function in vivo is protoplast-based transient expression analysis. We established a rice protoplast system about 10 years ago, which has been recently used in many laboratories. This protocol is useful for protein expression, subcellular localization, bimolecular fluorescence complementation, and co-immunoprecipitation assays. © 2016 by John Wiley & Sons, Inc.

Rice (Oryza sativa) is not only the staple food for half of the world's population but also a model monocot plant for molecular biology studies. Although rice genes have been extensively investigated in the last two decades, the functions of many genes in the rice genome are still not known. One of the rapid and efficient approaches for determining gene function in vivo is protoplast-based transient expression analysis. We established a rice protoplast system about 10 years ago, which has been recently used in many laboratories. This protocol is useful for protein expression, subcellular localization, bimolecular fluorescence complementation, and co-immunoprecipitation assays. © 2016 by John WileySons, Inc.

Background Rice (Oryza sativa) is the main food for half of the world’s population, and is considered the model for molecular biology studies of monocotyledon species. Although the rice genome was completely sequenced about 15 years ago, the function of most rice genes is still unknown. Results In this study, we developed a vector toolkit that contains 42 vectors for transient expression studies in rice protoplasts and stable expression analysis in transgenic rice. These vectors have been successfully used to study protein subcellular localization, protein-protein interaction, gene overexpression, and the CRISPR/Cas9-mediated gene editing. A novel feature of these vectors is that they contain a universal multiple cloning site, which enables more than 99% of the rice coding sequences to be conveniently transferred between vectors. Conclusions The versatile vectors represent a highly efficient and high-throughput toolkit for functional analysis of rice genes. Electronic supplementary material The online version of this article (10.1186/s12284-018-0220-7) contains supplementary material, which is available to authorized users.