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The economical production of 5‐aminolevulinic acid (ALA) has recently received increasing attention for its extensive use in agriculture. In this study, a strain of Bacillus cereus PT1 could initially produce ALA at a titre of 251.72 mg/L by using a hydrolysate mixture of low‐cost cassava residue and fish waste. The integration of endogenous hemA encoding glutamyl‐tRNA reductase led to a 39.30% increase in ALA production. Moreover, improving cell permeability by deletion of the LytR‐CpsA‐Psr (LCP) family gene tagU led to a further increase of 59.73% in ALA production. Finally, the engineered strain B. cereus PT1‐hemA‐ΔtagU produced 2.62 g/L of ALA from the previously mentioned hydrolysate mixture in a 7‐L bioreactor. In a pot experiment, foliar spray of the ALA produced by B. cereus PT1‐hemA‐ΔtagU from the hydrolysates increased salt tolerance of cucumber by improving chlorophyll content and catalase activity, while decreasing malondialdehyde content. Overall, this study demonstrated an economic way to produce ALA using a microbial platform and evidenced the potential of ALA in agricultural application. [ABSTRACT FROM AUTHOR]

A core-shell curing agent, in which E-44 epoxy resin modified 4,4'-diaminodiphenylmethane (M-DDM) is used as the core material and toluene-2,4-diisocyanate (TDI) as the shell monomer, was prepared by an in situ interfacial polymerization technique to prolong the shelf life of the composite repair prepreg system. The shell was formed on the surface of the M-DDM particles by the polyaddition of M-DDM and TDI monomers, and the mean thickness of the shell is about 100 nm. The core-shell particles showed an irregular sphere-like shape with a narrow size distribution, and the mean size of the core-shell particles was about 2.54 μm. The core-shell curing agent exhibited an excellent curing ability and a promising storage property of about 6 months, which can be applied in the field of the prepreg system for the rapid composite repair of the damaged structures. POLYM. COMPOS., 34:1892-1900, 2013. © 2013 Society of Plastics Engineers [ABSTRACT FROM AUTHOR]

SUMMARY The diversity in alternative splicing of long noncoding RNAs (lncRNAs) poses a challenge for functional annotation of lncRNAs. Moreover, little is known on the effects of alternatively spliced lncRNAs on crop yield. In this study, we cloned nine isoforms resulting from the alternative splicing of the lncRNA LAIR in rice. The LAIR isoforms are generated via alternative 5′/3′ splice sites and different combinations of specific introns. All LAIR isoforms activate the expression of the neighboring LRK1 gene and enhance yield‐related rice traits. In addition, there are slight differences in the binding ability of LAIR isoforms to the epigenetic modification‐related proteins OsMOF and OsWDR5, which affect the enrichment of H4K16ac and H3K4me3 at the LRK1 locus, and consequently fine‐tune the regulation of LRK1 expression and yield‐related traits. These differences in binding may be caused by polymorphic changes to the RNA secondary structure resulting from alternative splicing. It was also observed that the composition of LAIR isoforms was sensitive to abiotic stress. These findings suggest that the alternative splicing of LAIR leads to the formation of a functional transcript population that precisely regulates yield‐related gene expression, which may be relevant for phenotypic polymorphism‐based crop breeding under changing environmental conditions. [ABSTRACT FROM AUTHOR]

Symbiotic nitrogen fixation (SNF) is a crucial process for nitrogen geochemical cycling and plant‐microbe interactions. Water‐soluble humic acid (WSHM), an active component of soil humus, has been shown to promote SNF in the legume‐rhizobial symbiosis, but its molecular mechanism remains largely unknown. To reveal the SNF‐promoting mechanism, we conducted transcriptomic analysis on soybean treated with WSHM. Our findings revealed that up‐ and downregulated differentially expressed genes (DEGs) were mainly involved in plant cell‐wall/membrane formation and plant defence/immunity in the early stage, while the late stage was marked by the flavonoid synthesis and ethylene biosynthetic process. Further study on representative DEGs showed that WSHM could inhibit GmBAK1d‐mediated immunity and BR signalling, thereby promoting rhizobial colonisation, infection, and nodulation, while not favoring pathogenic bacteria colonisation on the host plant. Additionally, we also found that the ethylene pathway is necessary for promoting the soybean nodulation by WSHM. This study not only provides a significant advance in our understanding of the molecular mechanism of WSHM in promoting SNF, but also provides evidence of the beneficial interactions among the biostimulator, host plant, and soil microbes, which have not been previously reported. Summary Statement: Water‐soluble humic acid promoted soybean‐rhizobia symbiosis and nodulation by inhibiting host immunity and ethylene pathway, without sacrificing defence to the pathogenic bacteria. [ABSTRACT FROM AUTHOR]

RNA‐binding proteins (RBPs) are components of the post‐transcriptional regulatory system, but their regulatory effects on complex traits remain unknown. Using an integrated strategy involving map‐based cloning, functional characterizations, and transcriptomic and population genomic analyses, we revealed that RBP‐K (LOC_Os08g23120), RBP‐A (LOC_Os11g41890), and RBP‐J (LOC_Os10g33230) encode proteins that form an RBP‐A‐J‐K complex that negatively regulates rice yield‐related traits. Examinations of the RBP‐A‐J‐K complex indicated RBP‐K functions as a relatively non‐specific RBP chaperone that enables RBP‐A and RBP‐J to function normally. Additionally, RBP‐J most likely affects GA pathways, resulting in considerable increases in grain and panicle lengths, but decreases in grain width and thickness. In contrast, RBP‐A negatively regulates the expression of genes most likely involved in auxin‐regulated pathways controlling cell wall elongation and carbohydrate transport, with substantial effects on the rice grain filling process as well as grain length and weight. Evolutionarily, RBP‐K is relatively ancient and highly conserved, whereas RBP‐J and RBP‐A are more diverse. Thus, the RBP‐A‐J‐K complex may represent a typical functional model for many RBPs and protein complexes that function at transcriptional and post‐transcriptional levels in plants and animals for increased functional consistency, efficiency, and versatility, as well as increased evolutionary potential. Our results clearly demonstrate the importance of RBP‐mediated post‐transcriptional regulation for the diversity of complex traits. Furthermore, rice grain yield and quality may be enhanced by introducing various complete or partial loss‐of‐function mutations to specific RBP genes using clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR‐associated protein 9 technology and by exploiting desirable natural tri‐genic allelic combinations at the loci encoding the components of the RBP‐A‐J‐K complex through marker‐assisted selection. [ABSTRACT FROM AUTHOR]

Amylose content (AC) is the main factor determining the palatability, viscosity, transparency, and digestibility of rice (Oryza sativa) grains. AC in rice grains is mainly controlled by different alleles of the Waxy (Wx) gene. The AP2/EREBP transcription factor OsEBP89 interacts with the MYC‐like protein OsBP5 to synergistically regulate the expression of Wx. Here, we determined that the GLYCOGEN SYNTHASE KINASE 5 (OsGSK5, also named SHAGGY‐like kinase 41 [OsSK41]) inhibits the transcriptional activation activity of OsEBP89 in rice grains during amylose biosynthesis. The loss of OsSK41 function enhanced Wx expression and increased AC in rice grains. By contrast, the loss of function of OsEBP89 reduced Wx expression and decreased AC in rice grains. OsSK41 interacts with OsEBP89 and phosphorylates four of its sites (Thr‐28, Thr‐30, Ser‐238, and Thr‐257), which makes OsEBP89 unstable and attenuates its interaction with OsBP5. Wx promoter activity was relatively weak when regulated by the phosphomimic variant OsEBP89E–OsBP5 but relatively strong when regulated by the nonphosphorylatable variant OsEBP89A–OsBP5. Therefore, OsSK41‐mediated phosphorylation of OsEBP89 represents an additional layer of complexity in the regulation of amylose biosynthesis during rice grain development. In addition, our findings provide four possible sites for regulating rice grain AC via precise gene editing. [ABSTRACT FROM AUTHOR]

Auxin is an important phytohormone in plants, and auxin signaling pathways in rice play key roles in regulating its growth, development, and productivity. To investigate how rice grain yield traits are regulated by auxin signaling pathways and to facilitate their application in rice improvement, we validated the functional relationships among regulatory genes such as OsIAA10, OsSK41, and OsARF21 that are involved in one of the auxin (OsIAA10) signaling pathways. We assessed the phenotypic effects of these genes on several grain yield traits across two environments using knockout and/or overexpression transgenic lines. Based on the results, we constructed a model that showed how grain yield traits were regulated by OsIAA10 and OsTIR1, OsAFB2, and OsSK41 and OsmiR393 in the OsSK41‐OsIAA10‐OsARF module and by OsARF21 in the transcriptional regulation of downstream auxin response genes in the OsSK41‐OsIAA10‐OsARF module. The population genomic analyses revealed rich genetic diversity and the presence of major functional alleles at most of these loci in rice populations. The strong differentiation of many major alleles between Xian/indica and Geng/japonica subspecies and/or among modern varieties and landraces suggested that they contributed to improved productivity during evolution and breeding. We identified several important aspects associated with the genetic and molecular bases of rice grain and yield traits that were regulated by auxin signaling pathways. We also suggested rice auxin response factor (OsARF) activators as candidate target genes for improving specific target traits by overexpression and/or editing subspecies‐specific alleles and by searching and pyramiding the 'best' gene allelic combinations at multiple regulatory genes in auxin signaling pathways in rice breeding programs. [ABSTRACT FROM AUTHOR]

Xylanolytic enzymes, with both endo‐xylanase and arabinoxylan arabinofuranohydrolase (AXH) activities, are attractive for the economically feasible conversion of recalcitrant arabinoxylan. However, their characterization and utilization of these enzymes in biotechnological applications have been limited. Here, we characterize a novel bifunctional enzyme, rAbf43A, cloned from a bacterial consortium that exhibits AXH and endo‐xylanase activities. Hydrolytic pattern analyses revealed that the AXH activity belongs to AXHd3 because it attacked only the C(O)‐3‐linked arabinofuranosyl residues of double‐substituted xylopyranosyl units of arabinoxylan and arabinoxylan‐derived oligosaccharides, which are usually resistant to hydrolysis. The enzyme rAbf43A also liberated a series of xylo‐oligosaccharides (XOSs) from beechwood xylan, xylohexaose and xylopentaose, indicating that rAbf43A exhibited endo‐xylanase activity. Homology modelling based on AlphaFold2 and site‐directed mutagenesis identified three non‐catalytic residues (H161, A270 and L505) located in the substrate‐binding pocket essential for its dual‐functionality, while the mutation of A117 located in the −1 subsite to the proline residue only affected its endo‐xylanase activity. Additionally, rAbf43A showed significant synergistic action with the bifunctional xylanase/feruloyl esterase rXyn10A/Fae1A from the same bacterial consortium on insoluble wheat arabinoxylan and de‐starched wheat bran degradation. When rXyn10A/Fae1A was added to the rAbf43A pre‐hydrolyzed reactions, the amount of released reducing sugars, xylose and ferulic acid increased by 9.43% and 25.16%, 189.37% and 93.54%, 31.39% and 32.30%, respectively, in comparison with the sum of hydrolysis products released by each enzyme alone. The unique characteristics of rAbf43A position it as a promising candidate not only for designing high‐performance enzyme cocktails but also for investigating the structure–function relationship of GH43 multifunctional enzymes. [ABSTRACT FROM AUTHOR]

Summary: Gamma‐aminobutyric acid (GABA) and delta‐aminolevulinic acid (ALA), playing important roles in agriculture, medicine and other fields, are multifunctional non‐protein amino acids with similar and comparable properties and biosynthesis pathways. Recently, microbial synthesis has become an inevitable trend to produce GABA and ALA due to its green and sustainable characteristics. In addition, the development of metabolic engineering and synthetic biology has continuously accelerated and increased the GABA and ALA yield in microorganisms. Here, focusing on the current trends in metabolic engineering strategies for microbial synthesis of GABA and ALA, we analysed and compared the efficiency of various metabolic strategies in detail. Moreover, we provide the insights to meet challenges of realizing industrially competitive strains and highlight the future perspectives of GABA and ALA production. [ABSTRACT FROM AUTHOR]

Heading date is an important agronomic trait in rice (Oryza sativa L.). We previously fine‐mapped two complementary genes, Late Heading Date 1 and Late Heading Date 2 (LH2), that control the late heading trait in rice. Here, we cloned and analyzed the function of LH2. LH2 was fine‐mapped to a 53‐kb genomic region. Sequencing analysis revealed that there were differences in the coding sequence of LOC_Os08g07740 between the varieties 'Bo B' and 'Yuefeng B'. Therefore, LOC_Os08g07740 was identified as a candidate gene for LH2 and subsequently determined that LH2 is a Type 3 allele of Days To Heading 8. Transgenic plants of Yuefeng B carrying LH2 from Bo B significantly delayed the heading date under short‐day and long‐day conditions. A yeast two‐hybrid analysis revealed that LH2 physically interacted with Heme Activator Protein (HAP) 5A, ‐H, ‐I, ‐L, ‐K, and HAP5‐LIKE. The HAP5 domain of HAP5 family members has various functions. Further analysis revealed that LH2 represses photoperiodic flowering by controlling the expression of Early Heading Date 1, Heading date 3a, and Rice Flowering Locus T1 in rice. Core Ideas: LH2 delayed heading by 17.6 d in short‐day conditions and >32.7 d in long‐day conditions.LH2 physically interacted with HAP5A, ‐H, ‐I, ‐L, ‐K, and HAP5‐LIKE.The HAP5 domain of the HAP5 family exhibited functional differentiation.LH2 is a Type 3 allele of DTH8. [ABSTRACT FROM AUTHOR]

Leaf‐color mutants have been extensively studied in rice (Oryza sativa L.) and many corresponding genes have been identified and cloned. However, the mechanism of complex leaf‐color mutations requires further study. In this study, we obtained an introgression line (TIL22) from a set of introgression lines raised using African cultivated rice (O. glaberrima Steud.) as the donor parent and O. sativa subsp. indica Teqing as the recipient. Compared with Teqing, TIL22 showed a yellow‐green leaf phenotype at the seedling stage, but the leaves gradually changed to green after the five‐leaf stage. The photosynthetic pigment contents of yellow‐green leaves at the seedling stage were significantly reduced and chloroplast development was retarded compared with those of Teqing. Genetic analysis indicated that the introgression line phenotype was controlled by a single nuclear gene, temporarily designated YGL22. Map‐based cloning and sequence analysis suggested that the candidate gene was likely to be LOC_Os01g15390, which encodes a chloroplast protein. Real‐time quantitative polymerase chain reaction (qPCR) analysis revealed that the YGL22 transcript level was significantly lower in TIL22 than that of Teqing at the seedling stage. We generated the ygl22 knockout mutant using the CRISPR/Cas9 system. The ygl22 mutants showed a similar phenotype to that of TIL22 at the seedling stage, suggesting that YGL22 may be involved in the early development of chloroplasts in rice. In addition, plant height, number of panicles per plant, and grain yield per plant of ygl22 were not significantly affected. The results indicate that YGL22 can be used as a trait marker gene in hybrid rice production. [ABSTRACT FROM AUTHOR]

Heading date is one of the most important traits in rice and regulated by multiple genes. Common wild rice is the ancestor of Asian cultivated rice and harbours abundant genetic diversity. To use wild rice resource in rice breeding, a set of 154 introgression lines (ILs) covering 93% of the genome of Jinghong common wild rice was constructed in the background 'Teqing', using 208 simple sequence repeat markers evenly distributed on 12 chromosomes. Among the ILs, the line JIL64 displayed late heading independent of photoperiod. Genetic analysis using the two F2 populations crossed ''Teqing'/JIL64 and JIL64/'Teqing' revealed that late flowering was controlled by a recessive gene on chromosome 8 (designated early heading date 8, ehd8), and ehd8 was fine mapped to the 50‐kb region flanked by markers RM22221 and 64Indel4. Sequencing and qRT‐PCR demonstrated that LOC_Os08g01410 and LOC_Os08g01420 were deleted in JIL64 and may be associated with the late heading of Jinghong common wild rice. These findings lay a practical foundation for characterizing ehd8, and the ILs help to mine genes from Jinghong common wild rice. [ABSTRACT FROM AUTHOR]

Common wild rice ( Oryza rufipogon Griff.) is an important genetic reservoir for rice improvement. We investigated a quantitative trait locus (QTL), qGP5-1, which is related to plant height, leaf size and panicle architecture, using a set of introgression lines of O. rufipogon in the background of the Indica cultivar Guichao2 ( Oryza sativa L.). We cloned and characterized qGP5-1 and confirmed that the newly identified gene OsEBS (enhancing biomass and spikelet number) increased plant height, leaf size and spikelet number per panicle, leading to an increase in total grain yield per plant. Our results showed that the increased size of vegetative organs in OsEBS-expressed plants was enormously caused by increasing cell number. Sequence alignment showed that OsEBS protein contains a region with high similarity to the N-terminal conserved ATPase domain of Hsp70, but it lacks the C-terminal regions of the peptide-binding domain and the C-terminal lid. More results indicated that OsEBS gene did not have typical characteristics of Hsp70 in this study. Furthermore, Arabidopsis ( Arabidopsis thaliana) transformed with OsEBS showed a similar phenotype to OsEBS-transgenic rice, indicating a conserved function of OsEBS among plant species. Together, we report the cloning and characterization of OsEBS, a new QTL that controls rice biomass and spikelet number, through map-based cloning, and it may have utility in improving grain yield in rice. [ABSTRACT FROM AUTHOR]

With 4 figures and 1 table Abstract Oilseed rape ( Brassica napus L.) is the second most important oilseed crop in the world, and greater production of rape oil is required to satisfy the growing demand for it. We previously found that the cell size and yield of rice ( Oryza sativa) and cotton can be increased by over-expression of the RNA-binding domains (RRMs) of the flowering control locus A (FCA) protein. We also found that B. napus FCA-RRM2 shares a high level of sequence similarity with the RRM2 domains of other plants. We isolated B. napus FCA-RRM2 from variety No.1 'Nannongyou' of Brassica napus. Transformation experiments demonstrated that over-expression of B. napus FCA-RRM2 increases plant size, organ size, cell size, plant productivity and oil content in transgenic rape plants by down-regulating the cell-cycle-related gene cyclin-B2-1, a known activator of cyclin-dependent kinase 1 at the G2/M boundary. These results provide a practical approach for the genetic improvement of B. napus and possibly provide a model for investigating cell size regulation. [ABSTRACT FROM AUTHOR]

A dwarf mutant, designated LB4D, was obtained among the progeny of backcrosses to a wild rice introgression line. Genetic analysis of LB4D indicated that the dwarf phenotype was controlled by a single semidominant dwarfing gene, which was named LB4D. The mutants were categorized as dn‐type dwarf mutants according to the pattern of internode reduction. In addition, gibberellin (GA) response tests showed that LB4D plants were neither deficient nor insensitive to GA. This study found that tiller formation by LB4D plants was decreased by 40% compared with the wild type, in contrast to other dominant dwarf mutants that have been identified, indicating that a different dwarfing mechanism might be involved in the LB4D dominant mutant. The reduction of plant height in F1 plants ranged from 27.9% to 38.1% in different genetic backgrounds, showing that LB4D exerted a stronger dominant dwarfing effect. Using large F2 and F3 populations derived from a cross between heterozygous LB4D and the japonica cultivar Nipponbare, the LB4D gene was localized to a 46 kb region between the markers Indel 4 and Indel G on the short arm of chromosome 11, and four predicted genes were identified as candidates in the target region. [ABSTRACT FROM AUTHOR]

In rice ( Oryza sativa L.), the number of panicles, spikelets per panicle and grain weight are important components of grain yield. These characteristics are controlled by quantitative trait loci (QTLs) and are derived from variation inherent in crops. As a result of the complex genetic basis of these traits, only a few genes involved in their control have been cloned and characterized. We have previously map-cloned a gene cluster including eight leucine-rich repeat receptor-like kinase ( LRK) genes in Dongxiang wild rice ( Oryza rufipogon Griff.), which increased the grain yield by 16%. In the present study, we characterized the LRK1 gene, which was contained in the donor parent (Dongxiang wild rice) genome and absent from the recurrent parent genome (Guichao2, Oryza sativa L. ssp. indica). Our data showed that rice LRK1 is a plasma membrane protein expressed constitutively in leaves, young panicles, roots and culms. The over-expression of rice LRK1 results in increased panicles, spikelets per panicle, weight per grain and enhanced cellular proliferation, leading to a 27.09% increase in total grain yield per plant. The increased number of panicles and spikelets per panicle are associated with increased branch number. Our data suggest that rice LRK1 regulates rice branch number by enhancing cellular proliferation. The functional characterization of rice LRK1 facilitates an understanding of the mechanisms involved in cereal crop yield, and may have utility in improving grain yield in cereal crops. [ABSTRACT FROM AUTHOR]

A set of 148 F9 recombinant inbred lines (RILs) was developed from the cross of an indica cultivar 93-11 and japonica cultivar DT713, showing strong F1 heterosis. Subsequently, two backcross F1 (BCF1) populations were constructed by backcrossing these 148 RILs to two parents, 93-11 and DT713. These three related populations (281BCF1 lines, 148 RILs) were phenotyped for six yield-related traits in two locations. Significant inbreeding depression was detected in the population of RILS and a high level of heterosis was observed in the two BCF1 populations. A total of 42 main-effect quantitative trait loci (M-QTLs) and 109 epistatic effect QTL pairs (E-QTLs) were detected in the three related populations using the mixed model approach. By comparing the genetic effects of these QTLs detected in the RILs, BCF1 performance and mid-parental heterosis ( HMP), we found that, in both BCF1 populations, the QTLs detected could be classified into two predominant types: additive and over-dominant loci, which indicated that the additive and over-dominant effect were more important than complete or partially dominance for M-QTLs and E-QTLs. Further, we found that the E-QTLs detected collectively explained a larger portion of the total phenotypic variation than the M-QTLs in both RILs and BCF1 populations. All of these results suggest that additive and over-dominance resulting from epistatic loci might be the primary genetic basis of heterosis in rice. [ABSTRACT FROM AUTHOR]