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The Genomes Uncoupled 4 (GUN4) is one of the retrograde signaling genes in Arabidopsis and its orthologs have been identified in oxygenic phototrophic organisms from cyanobacterium to higher plants. GUN4 is involved in tetrapyrrole biosynthesis and its mutation often causes chlorophyll-deficient phenotypes with increased levels of reactive oxygen species (ROS), hence it has been speculated that GUN4 may also play a role in photoprotection. However, the biological mechanism leading to the increased ROS accumulation in gun4 mutants remains largely unknown. In our previous studies, we generated an epi-mutant allele of OsGUN4 (gun4epi), which downregulated its expression to ∼0.5% that of its wild-type (WT), and a complete knockout allele gun4-1 due to abolishment of its translation start site. In the present study, three types of F2 plant derived from a gun4-1/gun4epi cross, i.e., gun4-1/gun4-1, gun4-1/gun4epi and gun4epi/gun4epi were developed and used for further investigation by growing them under photoperiodic condition (16 h/8 h light/dark) with low light (LL, 100 μmol photons m–2 s–1) or high light (HL, 1000 μmol photons m–2 s–1). The expression of OsGUN4 was light responsive and had two peaks in the daytime. gun4-1/gun4-1-F2 seeds showed defective germination and died within 7 days. Significantly higher levels of ROS accumulated in all types of OsGUN4 mutants than in WT plants under both the LL and HL conditions. A comparative RNA-seq analysis of WT variety LTB and its gun4epi mutant HYB led to the identification of eight peroxidase (PRX)-encoding genes that were significantly downregulated in HYB. The transcription of these eight PRX genes was restored in transgenic HYB protoplasts overexpressing OsGUN4, while their expression was repressed in LTB protoplasts transformed with an OsGUN4 silencing vector. We conclude that OsGUN4 is indispensable for rice, its expression is light- and oxidative-stress responsive, and it plays a role in ROS accumulation via its involvement in the transcriptional regulation of PRX genes.

Abstract Background The cyclic nucleotide-gated ion channel (CNGC) family affects the uptake of cations, growth, pathogen defence, and thermotolerance in plants. However, the systematic identification, origin and function of this gene family has not been performed in Brassica oleracea, an important vegetable crop and genomic model organism. Results In present study, we identified 26 CNGC genes in B. oleracea genome, which are non-randomly localized on eight chromosomes, and classified into four major (I-IV) and two sub-groups (i.e., IV-a and IV-b). The BoCNGC family is asymmetrically fractioned into the following three sub-genomes: least fractionated (14 genes), most fractionated-I (10), and most fractionated-II (2). The syntenic map of BoCNGC genes exhibited strong relationships with the model Arabidopsis thaliana and B. rapa CNGC genes and provided markers for defining the regions of conserved synteny among the three genomes. Both whole-genome triplication along with segmental and tandem duplications contributed to the expansion of this gene family. We predicted the characteristics of BoCNGCs regarding exon-intron organisations, motif compositions and post-translational modifications, which diversified their structures and functions. Using orthologous Arabidopsis CNGCs as a reference, we found that most CNGCs were associated with various protein–protein interaction networks involving CNGCs and other signalling and stress related proteins. We revealed that five microRNAs (i.e., bol-miR5021, bol-miR838d, bol-miR414b, bol-miR4234, and bol-miR_new2) have target sites in nine BoCNGC genes. The BoCNGC genes were differentially expressed in seven B. oleracea tissues including leaf, stem, callus, silique, bud, root and flower. The transcript abundance levels quantified by qRT-PCR assays revealed that BoCNGC genes from phylogenetic Groups I and IV were particularly sensitive to cold stress and infections with bacterial pathogen Xanthomonas campestris pv. campestris, suggesting their importance in abiotic and biotic stress responses. Conclusion Our comprehensive genome-wide analysis represents a rich data resource for studying new plant gene families. Our data may also be useful for breeding new B. oleracea cultivars with improved productivity, quality, and stress resistance.

Kelch-like ECH-associated protein 1 (KEAP1)–nuclear factor E2-related factor 2 (NRF2) is the key antioxidant system in animals. In a previous study, we identified a probable KEAP1 ortholog in rice, OsKEAP1, and demonstrated that the downregulation of OsKEAP1 could alter the redox system and impair plant growth, as well as increase the susceptibility to abscisic acid (ABA) in seed germination. However, no NRF2 orthologs have been identified in plants and the mechanism underlying the phenotype changes of downregulated oskeap1 mutants is yet unknown. An in silico search showed that OsABI5 is the gene that encodes a protein with the highest amino acid identity score (38.78%) to NRF2 in rice. In this study, we demonstrated that, via yeast two-hybrids analysis and bimolecular fluorescence complementation assays, OsKEAP1 interacted with OsABI5 via its Kelch repeat domain in the nucleus. In germinating seeds, the expression of OsKEAP1 was significantly downregulated in oskeap1-1 (39.5% that of the wild-type (WT)) and oskeap1-2 (64.5% that of WT), while the expression of OsABI5 was significantly increased only in oskeap1-1 (247.4% that of WT) but not in oskeap1-2 (104.8% that of WT). ABA (0.5 μM) treatment significantly increased the expression of OsKEAP1 and OsABI5 in both the oskeap1 mutants and WT, and 4 days post treatment, the transcription level of OsABI5 became significantly greater in oskeap1-1 (+87.2%) and oskeap1-2 (+55.0%) than that in the WT. The ABA-responsive genes (OsRab16A and three late embryogenesis abundant genes), which are known to be activated by OsABI5, became more responsive to ABA in both oskeap1 mutants than in the WT. The transcript abundances of genes that regulate OsABI5, e.g., OsSnRK2 (encodes a kinase that activates OsABI5), OsABI1, and OsABI2 (both encode proteins binding to OsSnRK2 and are involved in ABA signaling) were not significantly different between the two oskeap1 mutants and the WT. These results demonstrated that OsKEAP1 played a role in the ABA response in rice seed germination via regulating OsABI5, which is the key player in the ABA response. In-depth analyses of the components and their action mode of the KEAP1–NRF2 and ABA signaling pathways suggested that OsKEAP1 likely formed a complex with OsABI5 and OsKEG, and OsABI5 was ubiquitinated by OsKEG and subsequently degraded under physiological conditions; meanwhile, under oxidative stress or with increased an ABA level, OsABI5 was released from the complex, phosphorylated, and transactivated the ABA response genes. Therefore, OsKEAP1–OsABI5 bore some resemblance to KEAP1–NRF2 in terms of its function and working mechanism.

Rice is one of the world’s most important crops, but its production suffers from insect pests. Rice brown planthopper (BPH; Nilaparvata lugens Stål) and striped stem borer (SSB, Chilo suppressalis Walker) are the two most serious pests in rice production. We reported that serotonin is an essential mediator in the interaction between rice and insect. Here, we established a method for extraction and determination of serotonin in rice and BPH.

The Kelch-like ECH-associated protein 1 (KEAP1)-nuclear factor E2-related factor 2 (NRF2) module is a key component in the detoxification and antioxidant system in animals, which plays crucial roles in cell homeostasis and cytoprotection, and consequently in carcinogenesis and disease development. However, this system seems to have diverged throughout evolution across different organisms, and the question of whether a similar system exists in plants has thus far remained unresolved. In this study, a KEAP1 ortholog was identified in rice (Oryza sativa L., OsKEAP1) and its properties were characterized via in silico and laboratory studies. To reveal OsKEAP1’s function, two knockdown mutants, oskeap1-1 and oskeap1-2, were generated by targeted mutagenesis in the 5′ untranslated region (UTR) using the CRISPR-Cas9 system. In silico analysis showed that OsKEAP1 has a Kelch-repeat domain which is identical to those of animals and a plant-specific development and cell death (DCD) domain in place of the broad-complex, tramtrack, bric-a-brac (BTB) domain found in animals. Orthologs of OsKEAP1 are present across plant species and all have the DCD domain and the Kelch-repeat domain. OsKEAP1 was proven to be localized to both the cytoplasm and nucleus, in contrast to the exclusive cytoplasm localization of animal KEAP1. Single nucleotide insertions in the 5′ UTR significantly reduced the transcription level of OsKEAP1 in the oskeap1-1 and oskeap1-2 mutants. The oskeap1 mutations greatly impaired plant growth and development, resulting in significant declines in a majority of agronomic and yield-related traits, i.e., plant height, panicle length, grain number per plant, and seed-set rate. The downregulation of OsKEAP1 increased the levels of H2O2, malondialdehyde, and proline while significantly decreasing the expression of two catalase genes in seedlings grown under normal and salt-stressed conditions. The changes in the above phenotypes are either positively or negatively correlated with the degree of OsKEAP1 downregulation. Altogether, we identified a probable KEAP1 ortholog in rice, revealed its unique subcellular localization, and demonstrated its important functions in vegetative and reproductive growth via regulation of the antioxidant response in plants.

The xantha marker trait, which is controlled by a down-regulating epi-mutation of OsGUN4, has been applied to the production of hybrid rice. However, the molecular basis for the ability of xantha mutants to attain high photosynthetic capacity even with decreased chlorophyll contents has not been characterized. In the present study, we observed that the total chlorophyll content of the xantha mutant was only 27.2% of that of the wild-type (WT) plants. However, the xantha mutant still accumulated 59.9% of the WT δ-aminolevulinic acid content, 72.8% of the WT Mg-protoporphyrin IX content, and 63.0% of the WT protochlorophyllide a content. Additionally, the protoporphyrin IX and heme contents in the mutant increased to 155.0 and 160.0%, respectively, of the WT levels. A search for homologs resulted in the identification of 124 rice genes involved in tetrapyrrole biosynthesis and photosynthesis. With the exception of OsGUN4, OsHO-1, and OsHO-2, the expression levels of the genes involved in tetrapyrrole biosynthesis were significantly higher in the xantha mutant than in the WT plants, as were all 72 photosynthesis-associated nuclear genes. In contrast, there were no differences between the xantha mutant and WT plants regarding the expression of all 22 photosynthesis-associated chloroplast genes. Furthermore, the abundance of 1O2 and the expression levels of 1O2-related genes were lower in the xantha mutant than in the WT plants, indicating 1O2-mediated retrograde signaling was repressed in the mutant plants. These results suggested that the abundance of protoporphyrin IX used for chlorophyll synthesis decreased in the mutant, which ultimately decreased the amount of chlorophyll in the xantha mutant. Additionally, the up-regulated expression of photosynthesis-associated nuclear genes enabled the mutant to attain a high photosynthetic capacity. Our findings confirm that OsGUN4 plays an important role in tetrapyrrole biosynthesis and photosynthesis in rice. GUN4, chlorophyll synthesis pathways, and photosynthetic activities are highly conserved in plants and hence, novel traits (e.g., xantha marker trait) may be generated in other cereal crops by modifying the GUN4 gene.

Nitrous oxide (N2O) is a potent greenhouse gas (GHG). Although it comprises only 0.03% of total GHGs produced, N2O makes a marked contribution to global warming. Much of the N2O in the atmosphere issues from incomplete bacterial denitrification processes acting on high levels of nitrogen (N) in the soil due to fertilizer usage. Using less fertilizer is the obvious solution for denitrification mitigation, but there is a significant drawback (especially where not enough N is available for the crop via N deposition, irrigation water, mineral soil N, or mineralization of organic matter): some crops require high-N fertilizer to produce the yields necessary to help feed the world’s increasing population. Alternatives for denitrification have considerable caveats. The long-standing promise of genetic modification for N fixation may be expanded now to enhance dissimilatory denitrification via genetic engineering. Biotechnology may solve what is thought to be a pivotal environmental challenge of the 21st century, reducing GHGs. Current approaches towards N2O mitigation are examined here, revealing an innovative solution for producing staple crops that can ‘crack’ N2O. The transfer of the bacterial nitrous oxide reductase gene (nosZ) into plants may herald the development of plants that express the nitrous oxide reductase enzyme (N2OR). This tactic would parallel the precedents of using the molecular toolkit innately offered by the soil microflora to reduce the environmental footprint of agriculture.

Background: The cyclic nucleotide-gated ion channel (CNGC) family affects the uptake of cations, growth, pathogen defence and thermotolerance in plants. However, the systematic identification, origin and function of this gene family has not been performed in Brassica oleracea, an important vegetable crop and genomic model organism. Results: In present study, we identified 26 CNGC genes in B. oleracea genome, which are non-randomly localized on eight chromosomes and classified into four major (I-IV) and two sub-groups (i.e., IV-a and IV-b). The BoCNGC family is asymmetrically fractioned into the following three sub-genomes: least fractionated (14 genes), most fractionated-I (10), and most fractionated-II (2). The syntenic map of BoCNGC genes exhibited strong relationships with the model Arabidopsis thaliana and B. rapa CNGC genes and provided markers for defining the regions of conserved synteny among the three genomes. Both whole-genome triplication along with segmental and tandem duplications contributed to the expansion of this gene family. We predicted the characteristics of BoCNGCs regarding exon-intron organisations, motif compositions and post-translational modifications, which diversified their structures and functions. Using orthologous Arabidopsis CNGCs as a reference, we found that most CNGCs were associated with various protein-protein interaction networks involving CNGCs and other signalling and stress related proteins. We revealed that five microRNAs (i.e., bol-miR5021, bol-miR838d, bol-miR414b, bol-miR4234 and bol-miR_new2) have target sites in nine BoCNGC genes. The BoCNGC genes were differentially expressed in seven B. oleracea tissues including leaf, stem, callus, silique, bud, root and flower. The transcript abundance levels quantified by qRT-PCR assays revealed that BoCNGC genes from phylogenetic Groups I and IV were particularly sensitive to cold stress and infections with bacterial pathogen Xanthomonas campestris pv. campestris, suggesting their importance in abiotic and biotic stress responses. Conclusion: Our comprehensive genome-wide analysis represents a rich data resource for studying new plant gene families. Our data may also be useful for breeding new B. oleracea cultivars with improved productivity, quality and stress resistance. [ABSTRACT FROM AUTHOR]

The recessive mutation of the XANTHA gene ( XNT) transforms seedlings and plants into a yellow color, visually distinguishable from normal (green) rice. Thus, it has been introduced into male sterile lines as a distinct marker for rapidly testing and efficiently increasing varietal purity in seed and paddy production of hybrid rice. To identify closely linked markers and eventually isolate the XNT gene, two mapping populations were developed by crossing the xantha mutant line Huangyu B ( indica) with two wild type japonica varieties; a total of 1,720 mutant type F2 individuals were analyzed for fine mapping using polymorphic InDel markers and high dense microsatellite markers. The XNT gene was mapped on chromosome 11, within in a fragment of ~100 kb, where 13 genes are annotated. The NP_001067671.1 gene within the delimited region is likely to be a candidate XNT gene, since it encodes ATP-dependent chloroplast protease ATP-binding subunit clp A. However, no sequence differences were observed between the mutant and its parent. Bioinformatics analysis demonstrated that four chlorophyll deficient mutations that were previously mapped on the same chromosome are located outside the XNT region, indicating XNT is a new gene. The results provide useful DNA markers not only for marker assisted selection of the xantha trait but also its eventual cloning. [ABSTRACT FROM AUTHOR]

One of the key steps in positional cloning and marker-aided selection is to identify marker(s) tightly linked to the target gene (i.e., fine mapping). Selective genotyping such as selective recombinant genotyping (SRG) is commonly used in fine mapping for cost-saving. To further decrease genotyping effort and rapidly screen for tightly linked markers, we propose here a combined DNA pooling and SRG strategy. A two-stage pooled genotyping can be used for identifying recombinants between a pair of flanking markers more efficiently, and a joint use of bulked DNA analysis and two-stage pooling can also save cost for genotyping recombinants. The combined DNA pooling and SRG strategy can further be extended to fine mapping for polygenic traits. The numerical results based on hypothetical scenarios and an illustrative application to fine mapping of a mutant gene, called xl( t), in rice suggest that the proposed strategy can remarkably reduce genotyping amount compared with the conventional SRG. [ABSTRACT FROM AUTHOR]

Phytic acid (PA, myo-inositol 1,2,3,4,5,6-hexa kisphosphate) is important to the nutritional quality of cereal and legume seeds. PA and its salts with micronutrient cations, such as iron and zinc, cannot be digested by humans and non-ruminant animals, and hence may affect food/feed nutritional value and cause P pollution of groundwater from animal waste. We previously developed a set of low phytic acid (LPA) rice mutant lines with the aim of increasing the nutritional quality of rice. Two of these lines, Os-lpa-XS110-2 (homozygous non-lethal) Os-lpa-XS110-3 (homozygous lethal), contain two mutant alleles of a LPA gene (hereafter XS-lpa2-1 and XS-lpa2-2, respectively). In this study, we mapped the XS-lpa2-1 gene to a region on chromosome 3 between microsatellite markers RM14360 and RM1332, where the rice orthologue ( OsMRP5) of the maize lpa1 gene is located. Sequence analysis of the OsMRP5 gene revealed a single base pair change (C/G–T/A transition) in the sixth exon of XS-lpa2-1 and a 5-bp deletion in the first exon of XS-lpa2-2. OsMRP5 is expressed in both vegetative tissues and developing seeds, and the two mutations do not change the level of RNA transcription. A T-DNA insertion line, 4A-02500, in which OsMRP5 was disrupted, also showed the same high inorganic phosphorus phenotype as Os-lpa-XS110-3 and appeared to be homozygous lethal. PA is significantly reduced in Os-lpa-XS110-2 (~20%) and in 4A-02500 (~90%) seeds compared with their wild type lines, and no PA was detected in Os-lpa-XS110-3 using HPLC analysis. This evidence indicates that the OsMRP5 gene plays an important role in PA metabolism in rice seeds. [ABSTRACT FROM AUTHOR]

Phytic acid (PA, myo-inositol 1,2,3,4,5,6-hexa kisphosphate) is an important anti-nutritional component in cereal and legume grains. PA forms of phosphorus (P) and its salts with micronutrient cations, such as iron and zinc, are indigestible in humans and non-ruminant animals, and hence could affect food/feed nutritional value and cause P pollution of ground water from animal waste. We previously developed a set of low phytic acid (LPA) rice mutants with the aim to increase their nutritional quality. Among them, one line, i.e., Os-lpa-XQZ-1 (hereafter lpa 1- 2), was identified to have a mutation allelic to the KBNT lpa 1- 1 mutation (hereafter lpa 1- 1), which was already delimited to a 47-kb region on chromosome 2. In this study, we searched the candidate gene for these two allelic LPA mutations using T-DNA insertion mutants, mutation detection by CEL I facilitated mismatch cleavage, and gene sequencing. The TIGR locus LOC_Os02g57400 was revealed as the candidate gene hosting these two mutations. Sequence analysis showed that the lpa 1- 1 is a single base pair substitution mutation, while lpa 1- 2 involves a 1,475-bp fragment deletion. A CAPS marker (LPA1_CAPS) was developed for distinguishing the lpa 1- 1 allele from lpa 1- 2 and WT alleles, and InDel marker (LPA1_InDel) was developed for differentiating the lpa 1- 2 allele from lpa 1- 1 and WT ones. Analysis of two populations derived from the two mutants with wild-type varieties confirmed the complete co-segregation of these two markers and LPA phenotype. The LOC_Os02g57400 is predicted to encode, through alternative splicing, four possible proteins that are homologous to the 2-phosphoglycerate kinase reported in hyperthermophilic and thermophilic bacteria. The identification of the LPA gene and development of allele-specific markers are of importance not only for breeding LPA varieties, but also for advancing genetics and genomics of phytic acid biosynthesis in rice and other plant species. [ABSTRACT FROM AUTHOR]

Mutation techniques have been used for generating genetic variation and breeding new varieties during the past decades. However, the skepticism has also persisted during the course on the sole mutational origin of genetic variation in mutated populations. We addressed this issue using three unique rice genetic lines in this study. First, we confirmed that gamma rays had significant effect on the growth of M1 plants, leading to significant reduction of fertility, seed set and plant height at doses 200 Gy and above. Second, we proved that out-crossing derived genetic variants existed in M2 population (0.8%) and among selected putative mutants (0–33.3%), in addition to induced mutants. Third, we demonstrated that true induced mutant lines had identical microsatellite haplotypes to their parents. We proposed microsatellite assay as a method to exclude any genetic contaminants from induced mutants, with appropriate numbers for different levels of power based on reported microsatellite mutation rate and microsatellite polymorphic index. [ABSTRACT FROM AUTHOR]

Phytic acid (PA, myo-inositol 1, 2, 3, 4, 5, 6 hexa kisphosphate) is important to the nutritional quality of soybean meal. Organic phosphorus (P) in PA is indigestible in humans and non-ruminant animals, which affects nutrition and causes P pollution of ground water from animal wastes. Two novel soybean [( Glycine max L. (Merr.)] low phytic acid ( lpa) mutations were isolated and characterized. Gm-lpa-TW-1 had a phytic acid P (PA-P) reduction of 66.6% and a sixfold increase in inorganic P (Pi), and Gm- lpa-ZC-2 had a PA-P reduction of 46.3% and a 1.4-fold increase in Pi, compared with their respective non-mutant progenitor lines. The reduction of PA-P and increase of Pi in Gm-lpa-TW-1 were molar equivalent; the decrease of PA-P in Gm- lpa-ZC-2, however, was accompanied by the increase of both Pi and lower inositol phosphates. In both mutant lines, the total P content remained similar to their wild type parents. The two lpa mutations were both inherited in a single recessive gene model but were non-allelic. Sequence data and progeny analysis indicate that Gm-lpa-TW-1 lpa mutation resulted from a 2 bp deletion in the soybean d- myo-inositol 3-phosphate synthase (MIPS1 EC 5.5.1.4) gene 1 ( MIPS1). The lpa mutation in Gm- lpa-ZC-2 was mapped on LG B2, closely linked with microsatellite loci Satt416 and Satt168, at genetic distances of ∼4.63 and ∼9.25 cM, respectively. Thus this mutation probably represents a novel soybean lpa locus. The seed emergence rate of Gm-lpa-ZC-2 was similar to its progenitor line and was not affected by seed source and its lpa mutation. However, Gm-lpa-TW-1 had a significantly reduced field emergence when seeds were produced in a subtropic environment. Field tests of the mutants and their progenies further demonstrated that the lpa mutation in Gm-lpa-ZC-2 does not negatively affect plant yield traits. These results will advance understanding of the genetic, biochemical and molecular control of PA synthesis in soybean. The novel lpa mutation in Gm-lpa-ZC-2, together with linked simple sequence repeat (SSR) markers, will be of value for breeding productive lpa soybeans, with meal high in digestible Pi eventually to improve animal nutrition and lessen environmental pollution. [ABSTRACT FROM AUTHOR]

Efficient mutant production systems, through either physical or chemical mutagenesis, have been well established in soybean. A vast amount of genetic variability, of both quantitative and qualitative traits, has been generated through experimental mutagenesis in the past 30 years. Characterization of mutated traits has greatly advanced our understanding of the underlying mechanisms of a score of important traits, including the genetic and genomic basis of nodulation and synthesis of fatty acids and storage proteins. Mutation techniques have also been successfully used in breeding new soybean varieties. More than 100 mutant varieties were developed and released for commercial cultivation; among them are several groundbreaking varieties; i.e., the supernodulating variety Sakukei 4; the glycinin-rich variety Yumenori; the lipoxygenases-free variety Ichihime; and the low linolenic varieties IA3017 and IA2064. It is expected that more new varieties with novel traits will become available in the coming years, and a variety with a reduced phytate level may become available in the near future. Induced mutants will become a unique genetic resource in functional genomic studies, and the advance of functional genomics will increase the usefulness and use efficiency of induced mutations in soybean breeding. [ABSTRACT FROM AUTHOR]

We previously demonstrated that mutants induced from pure lines with quality assurance had microsatellite (or simple sequence repeat, SSR) genotypes almost identical to their parent varieties, and genetic variants derived from accidental out-crossing might be selected as "mutants" in rice. In the present study, fourteen previously developed rice mutant lines were analyzed for their SSR genotypes in parallel with their respective parents. Among them, eleven mutant lines had SSR genotypes identical to their parents; however, three mutant lines, i.e., Os-lpa-XS110-1, Huangyu B, and Zhu I, had 4.3, 6.6, and 9.1% of SSRs different from their parents, respectively. Additional analysis proved that Huangyu B was mistakenly recorded in the development process and should be a mutant line of another variety, II 32 B; Zhu I very probably resulted from an out-crossing rather than from mutagenesis. The implications of our findings are discussed in the context of induced mutantfacilitated gene discovery. [ABSTRACT FROM AUTHOR]

Phytic acid (PA, myo-inositol 1,2,3,4,5,6-hexa kisphosphate), or its salt form, phytate, is commonly regarded as the major anti-nutritional component in cereal and legume grains. Breeding of low phytic acid ( lpa) crops has recently been considered as a potential way to increase nutritional quality of crop products. In this study, eight independent lpa rice mutant lines from both indica and japonica subspecies were developed through physical and chemical mutagenesis. Among them, five are non-lethal while the other three are homozygous lethal. None of the lethal lines could produce homozygous lpa plants through seed germination and growth under field conditions, but two of them could be rescued through in vitro culture of mature embryos. The non-lethal lpa mutants had lower PA content ranging from 34 to 64% that of their corresponding parent and four of them had an unchanged total P level. All the lpa mutations were inherited in a single recessive gene model and at least four lpa mutations were identified mutually non-allelic, while the other two remain to be verified. One mutation was mapped on chromosome 2 between microsatellite locus RM3542 and RM482, falling in the same region as the previously mapped lpa1-1 locus did; another lpa mutation was mapped on chromosome 3, tightly linked to RM3199 with a genetic distance of 1.198 cM. The latter mutation was very likely to have happened to the LOC_Os03g52760, a homolog of the maize myo-inositol kinase (EC 2.7.1.64) gene. The present work greatly expands the number of loci that could influence the biosynthesis of PA in rice, making rice an excellent model system for research in this area. [ABSTRACT FROM AUTHOR]

Comparative analysis revealed that a xantha rice mutant (cv. Huangyu B) had higher ratios of chlorophyll (Chl) a/b and carotenoids/Chl, and higher photosynthetic efficiency than its wild type parent (cv. II32 B). Unexpectedly, the mutant had higher net photosynthetic rate (PN) than II32 B. This might have resulted from its lower non-photochemical quenching (qN) but higher maximal photochemical efficiency (FV/FM), higher excitation energy capture efficiency of photosystem 2 (PS2) reaction centres (FV′/FM′), higher photochemical quenching (qP), higher effective PS2 quantum yield (ΦPS2), and higher non-cyclic electron transport rate (ETR). This is the first report of a chlorophyll mutant that has higher photosynthetic efficiency and main Chl fluorescence parameters than its wild type. This mutant could become a unique material both for the basic research on photosynthesis and for the development of high yielding rice cultivars. [ABSTRACT FROM AUTHOR]

The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated (Cas) technology has recently emerged as a powerful genomic editing tool with great potential for crop breeding. However, commonly used protocols for screening of CRISPR/Cas9-induced mutations are laborious, timeconsuming, and costly. In the present study we examined the applicability of high resolution melting (HRM) analysis fast screening CRISPR/Cas9-induced mutations in T0 plants and subsequent genotyping of T1 populations. Comparative analysis demonstrated that HRM analysis could identify mutant T0 plants carrying various types of mutation, including single nucleotide substitutions and short insertion/deletions, with false positive/negative rates of 0-2.78%. Furthermore, T1 plants derived from single T0 plants could be correctly genotyped by HRM analysis using WT parents and T0 plants as controls. We hence recommend the adoption of HRM analysis in CRISPR/Cas9 mediated genetic studies and breeding in rice and other crop species. [ABSTRACT FROM AUTHOR]

Doubled haploid (DH) populations, particularly those from subspecies crosses possessing the wide compatible gene S5n, are important germplasm resources for rice genetic studies and breeding, but their feature and potential have not been fully assessed and explored. In the present study, we produced a DH population from the hybrid of japonica 668B and wide compatible indica T23. Genotyping of the S5 locus with allele-specific markers for ORF3, ORF4 and ORF5 revealed a potential recombination hot spot in the ORF3-ORF4 region. Haplotyping analysis revealed that 21/34 subspecies specific Indel markers segregated in distortion in the DH population, with a few lines having indica alleles either extremely low (1.7%) or high (98.3%), with little effect of the S5 allele. While DH lines with the S5n allele had higher frequency of indica alleles, no effect of the S5n allele was observed on all agronomic traits but flowering time. Taken together, the present study advanced understanding of the genetics of wide crosses in general, and DH production in particular between the two rice subspecies, and the new DH population generated will become a useful resource for rice genetic study and breeding in the future. [ABSTRACT FROM AUTHOR]