Graphene oxide and indole-3-acetic acid cotreatment regulates the root growth of Brassica napus L. via multiple phytohormone pathways.

BACKGROUND: Studies have indicated that graphene oxide (GO) could regulated Brassica napus L. root growth via abscisic acid (ABA) and indole-3-acetic acid (IAA). To study the mechanism and interaction between GO and IAA further, B. napus L (Zhongshuang No. 9) seedlings were treated with GO and IAA accordance with a two factor completely randomized design. RESULTS: GO and IAA cotreatment significantly regulated the root length, number of adventitious roots, and contents of IAA, cytokinin (CTK) and ABA. Treatment with 25 mg/L GO alone or IAA (> 0.5 mg/L) inhibited root development. IAA cotreatment enhanced the inhibitory role of GO, and the inhibition was strengthened with increased in IAA concentration. GO treatments caused oxidative stress in the plants. The ABA and CTK contents decreased; however, the IAA and gibberellin (GA) contents first increased but then decreased with increasing IAA concentration when IAA was combined with GO compared with GO alone. The 9-cis-epoxycarotenoid dioxygenase (NCED) transcript level strongly increased when the plants were treated with GO. However, the NCED transcript level and ABA concentration gradually decreased with increasing IAA concentration under GO and IAA cotreatment. GO treatments decreased the transcript abundance of steroid 5-alpha-reductase (DET2) and isochorismate synthase 1 (ICS), which are associated with brassinolide (BR) and salicylic acid (SA) biosynthesis, but increased the transcript abundance of brassinosteroid insensitive 1-associated receptor kinase 1 (BAK1), cam-binding protein 60-like G (CBP60) and calmodulin binding protein-like protein 1, which are associated with BR and SA biosynthesis. Last, GO treatment increased the transcript abundance of 1-aminocyclopropane-1-carboxylic acid synthase 2 (ACS2), which is associated with the ethylene (ETH) pathway. CONCLUSIONS: Treatment with 25 mg/L GO or IAA (> 0.5 mg/L) inhibited root development. However, IAA and GO cotreatment enhanced the inhibitory role of GO, and this inhibition was strengthened with increased IAA concentration. IAA is a key factor in the response of B. napus L to GO and the responses of B. napus to GO and IAA cotreatment involved in multiple pathways, including those involving ABA, IAA, GA, CTK, BR, SA. Specifically, GO and IAA cotreatment affected the GA content in the modulation of B. napus root growth.

A metabolome-based core hybridisation strategy for the prediction of rice grain weight across environments.

Marker-based prediction holds great promise for improving current plant and animal breeding efficiencies. However, the predictabilities of complex traits are always severely affected by negative factors, including distant relatedness, environmental discrepancies, unknown population structures, and indeterminate numbers of predictive variables. In this study, we utilised two independent F1 hybrid populations in the years 2012 and 2015 to predict rice thousand grain weight (TGW) using parental untargeted metabolite profiles with a partial least squares regression method. A stable predictive model for TGW was built based on hybrids from the population in 2012 (r = 0.75) but failed to properly predict TGW for hybrids from the population in 2015 (r = 0.27). After integrating hybrids from both populations into the training set, the TGW of hybrids could be predicted but was largely dependent on population structures. Then, core hybrids from each population were determined by principal component analysis and the TGW of hybrids in both environments were successfully predicted (r > 0.60). Moreover, adjusting the population structures and numbers of predictive analytes increased TGW predictability for hybrids in 2015 (r = 0.72). Our study demonstrates that the TGW of F1 hybrids across environments can be accurately predicted based on parental untargeted metabolite profiles with a core hybridisation strategy in rice. Metabolic biomarkers identified from early developmental stage tissues, which are grown under experimental conditions, may represent a workable approach towards the robust prediction of major agronomic traits for climate-adaptive varieties.

Rice PPS1 encodes a DYW motif-containing pentatricopeptide repeat protein required for five consecutive RNA-editing sites of nad3 in mitochondria.

The pentatricopeptide repeat (PPR) protein family is a large family characterized by tandem arrays of a degenerate 35-amino-acid motif whose members function as important regulators of organelle gene expression at the post-transcriptional level. Despite the roles of PPRs in RNA editing in organelles, their editing activities and the underlying mechanism remain obscure. Here, we show that a novel DYW motif-containing PPR protein, PPS1, is associated with five conserved RNA-editing sites of nad3 located in close proximity to each other in mitochondria, all of which involve conversion from proline to leucine in rice. Both pps1 RNAi and heterozygous plants are characterized by delayed development and partial pollen sterility at vegetative stages and reproductive stage. RNA electrophoresis mobility shift assays (REMSAs) and reciprocal competition assays using different versions of nad3 probes confirm that PPS1 can bind to cis-elements near the five affected sites, which is distinct from the existing mode of PPR-RNA binding because of the continuity of the editing sites. Loss of editing at nad3 in pps1 reduces the activity of several complexes in the mitochondrial electron transport chain and affects mitochondrial morphology. Taken together, our results indicate that PPS1 is required for specific editing sites in nad3 in rice.

Gnp4/LAX2, a RAWUL protein, interferes with the OsIAA3-OsARF25 interaction to regulate grain length via the auxin signaling pathway in rice.

Grain length is one of the determinants of yield in rice and auxin plays an important role in regulating it by mediating cell growth. Although several genes in the auxin pathway are involved in regulating grain length, the underlying molecular mechanisms remain unclear. In this study we identify a RING-finger and wd40-associated ubiquitin-like (RAWUL) domain-containing protein, Gnp4/LAX2, with a hitherto unknown role in regulation of grain length by its influence on cell expansion. Gnp4/LAX2 is broadly expressed in the plant and subcellular localization analysis shows that it encodes a nuclear protein. Overexpression of Gnp4/LAX2 can significantly increase grain length and thousand-kernel weight. Moreover, Gnp4/LAX2 physically interacts with OsIAA3 and consequently interferes with the OsIAA3-OsARF25 interaction in vitro and in vivo. OsIAA3 RNAi plants consistently exhibit longer grains, while the mutant osarf25 has small grains. In addition, OsARF25 binds to the promoter of OsERF142/SMOS1, a regulator of organ size, and positively regulates its expression. Taken together, the results reveal that Gnp4/LAX2 functions as a regulator of grain length through participation in the OsIAA3-OsARF25-OsERF142 pathway and that it has potential value for molecular breeding in rice.

Graphene Oxide Affects Growth and Resistance to Sclerotinia sclerotiorum in Brassica napus L.

Researchers have shown that graphene oxide has a significant effect on plant and pathogen growth and development. To better understand the effect of graphene oxide on the resistance of Brassica napus L. to Sclerotinia sclerotiorum, Zhongshuang 11 was used to evaluate changes in the morphology and physiology after graphene oxide treatment. Detached leaf inoculation was used to detect S. sclerotiorum infection in rapeseed. The results indicated that treatment with a low concentration of graphene oxide had no significant effect on the growth of B. napus "Zhongshuang 11." Graphene oxide inhibited S. sclerotiorum in PDA medium. Treatment with 15 mg/L graphene oxide for 8-24 h in seeds and 8-16 h in seedlings suppressed S. sclerotiorum growth compared to the control samples. These results demonstrate that a low concentration of graphene oxide did not harm the growth of B. napus but did enhance its resistance to S. sclerotiorum.

Pentatricopeptide-repeat family protein RF6 functions with hexokinase 6 to rescue rice cytoplasmic male sterility.

Cytoplasmic male sterility (CMS) has been extensively used for hybrid seed production in many major crops. Honglian CMS (HL-CMS) is one of the three major types of CMS in rice and has contributed greatly to food security worldwide. The HL-CMS trait is associated with an aberrant chimeric mitochondrial transcript, atp6-orfH79, which causes pollen sterility and can be rescued by two nonallelic restorer-of-fertility (Rf) genes, Rf5 or Rf6. Here, we report the identification of Rf6, which encodes a novel pentatricopeptide repeat (PPR) family protein with a characteristic duplication of PPR motifs 3-5. RF6 is targeted to mitochondria, where it physically associates with hexokinase 6 (OsHXK6) and promotes the processing of the aberrant CMS-associated transcript atp6-orfH79 at nucleotide 1238, which ensures normal pollen development and restores fertility. The duplicated motif 3 of RF6 is essential for RF6-OsHXK6 interactions, processing of the aberrant transcript, and restoration of fertility. Furthermore, reductions in the level of OsHXK6 result in atp6-orfH79 transcript accumulation and male sterility. Together these results reveal a novel mechanism for CMS restoration by which RF6 functions with OsHXK6 to restore HL-CMS fertility. The present study also provides insight into the function of hexokinase 6 in regulating mitochondrial RNA metabolism and may facilitate further exploitation of heterosis in rice.

OsLG3 contributing to rice grain length and yield was mined by Ho-LAMap.

BACKGROUND: Most agronomic traits in rice are complex and polygenic. The identification of quantitative trait loci (QTL) for grain length is an important objective of rice genetic research and breeding programs. RESULTS: Herein, we identified 99 QTL for grain length by GWAS based on approximately 10 million single nucleotide polymorphisms from 504 cultivated rice accessions (Oryza sativa L.), 13 of which were validated by four linkage populations and 92 were new loci for grain length. We scanned the Ho (observed heterozygosity per locus) index of coupled-parents of crosses mapping the same QTL, based on linkage and association mapping, and identified two new genes for grain length. We named this approach as Ho-LAMap. A simulation study of six known genes showed that Ho-LAMap could mine genes rapidly across a wide range of experimental variables using deep-sequencing data. We used Ho-LAMap to clone a new gene, OsLG3, as a positive regulator of grain length, which could improve rice yield without influencing grain quality. Sequencing of the promoter region in 283 rice accessions from a wide geographic range identified four haplotypes that seem to be associated with grain length. Further analysis showed that OsLG3 alleles in the indica and japonica evolved independently from distinct ancestors and low nucleotide diversity of OsLG3 in indica indicated artificial selection. Phylogenetic analysis showed that OsLG3 might have much potential value for improvement of grain length in japonica breeding. CONCLUSIONS: The results demonstrated that Ho-LAMap is a potential approach for gene discovery and OsLG3 is a promising gene to be utilized in genomic assisted breeding for rice cultivar improvement.

Interaction with Cu²âº disrupts the RNA binding affinities of RNA recognition motif containing protein.

The glycine-rich proteins (GRP) containing RNA recognition motifs (RRM) are involved in the regulation of transcriptional and/or post-transcriptional events. Previous studies have established that GRP162 plays an important role in the restoration of fertility in Honglian cytoplasmic male sterile (HL-CMS) rice. In this study, the ion binding properties of rGRP162 were tested by isothermal titration calorimetry (ITC) and electrophoretic mobility shift assay (EMSA) was performed to test the interaction. Circular dichroism (CD) was carried out to detect the alteration of secondary structure in the presence and absence of Cu(2+). Furthermore, two RRM containing proteins, AtRBP45A and AtRBP47A, were expressed to validate the interaction. Results showed Cu(2+) and Fe(3+) bound GRP162, whereas Ca(2+), Mn(2+), Mg(2+) and K(+) did not. EMSA confirmed that interaction with Cu(2+) interrupted the biological activity of GRP162 by disrupting the secondary structure of the protein based on the results of CD. Moreover, the RNA binding activities of rAtRBP45A and rAtRBP47A were also impaired in the presence of Cu(2+). Data suggest that Cu(2+) in excess may disrupt RNA-binding proteins containing RRM that are essential for post-transcriptional regulation and may impair the development of plants or animals.

Preparation of Antimony-Doped Tin Oxide Fly Ash Antistatic Composite and Its Properties in Filling EVA.

As a common coal-based solid waste, fly ash is widely used in material filling. However, due to the high resistivity of fly ash itself, the antistatic performance of the filling material is poor. Therefore, antistatic composite powder was prepared by coating nano-sized antimony-doped tin oxide (ATO) on the surface of fly ash, and its preparation mechanism was discussed. The composite powders were characterized by SEM, EDS, XRD and FTIR. The results show that the interaction between SiO2 and SnO2 appears at the wave number of 727.12 cm-1, and the obvious SnO2 crystal phase appears on the surface of fly ash. The volume resistivity of calcined fly ash is 1.72 × 1012 Ω·cm, and the volume resistivity of ATO fly ash is reduced to 6 × 103 Ω·cm. By analyzing the limiting oxygen index, melt index, tensile strength, elongation at break, cross-section morphology and surface electrical resistivity of EVA, it was found that the addition of antistatic powder to EVA can improve its antistatic performance without deteriorating the mechanical properties of EVA.

The mechanism of ORFH79 suppression with the artificial restorer fertility gene Mt-GRP162.

The restoration fertility complex (RFC) was previously identified in Honglian (HL)-cytoplasmic male sterility (CMS) rice (Oryza sativa), and glycine-rich protein 162 (GRP162) is responsible for binding to the CMS-associated transcript atp6-orfH79. Here, we engineered a recombinant GRP162 containing the mitochondrial transit peptide, termed Mt-GRP162, as an artificial restorer of fertility (Rf) gene. Mt-GRP162 was confirmed to bind to CMS-associated RNA and to localize to the mitochondria. The transgenic plants showed restored fertility with partially functional pollen. We found that the expression of ORFH79 decreased in transgenic plants, while the expression of atp6-orfH79 was not changed. These findings indicate that Mt-GRP162 restores fertility by suppressing the expression of the cytotoxic protein ORFH79 at the post-transcriptional level rather than via the cleavage of atp6-orfH79 in the presence of RFC. These findings contribute to our understanding of the mechanisms of restoration through diverse pathways.

Characterization of the complete chloroplast genome of the Pistacia vera L.

The complete chloroplast genome sequence of Pistacia vera was mapped and determined based on Illumina sequencing data. The complete chloroplast genome is 160,654 bp and contains a pair of inverted repeat regions of 26,596 bp each, a large single-copy region of 88,376 bp, and a small single-copy region of 19,086 bp. It harbors 113 genes, including 79 protein-coding genes,4 ribosomal RNA genes, and 30 transfer RNA genes. Phylogenetic analysis based on chloroplast genomes indicates that Pistacia vera is closely related to that of Pistacia weinmanniifolia.

Graphene oxide and ABA cotreatment regulates root growth of Brassica napus L. by regulating IAA/ABA.

Previous studies have proven that graphene oxide (GO) regulates abscisic acid (ABA) and indole-3-acetic acid (IAA) contents and modulates plant root growth. To better understand the mechanism of plant growth and development regulated by GO and crosstalk between ABA and GO, Zhongshuang No. 9 seedlings were treated with GO and ABA. The results indicated that GO and ABA significantly affected the morphological properties and endogenous phytohormone contents in seedlings, and there was significant crosstalk between GO and ABA. ABA treatments combined with GO led to a rapid decrease in triphenyltetrazolium chloride (TTC) reduction intensity, and the inhibitory effect was enhanced with increasing ABA concentration. The treatments significantly affected the transcriptional levels of some key genes involved in the ABA, IAA, cytokinin (CTK), salicylic acid (SA), and ethane (ETH) pathways and increased the ABA and gibberellin (GA) contents in rapeseed seedlings. The effects of the treatments on the IAA and CTK contents were complex, but, importantly, the treatments suppressed root elongation. Correlation analysis also indicated that the relationship between root length and IAA/ABA could be described by a polynomial function: y = 88.11x2 - 25.15x + 4.813(R²â€¯= 0.912). The treatments increased the ACS2 transcript abundance for ETH biosynthesis and the ICS1 transcriptional level of the key genes involved in salicylic acid (SA) biosynthesis, as well as the downstream signaling genes CBP60 and SARD1. This finding indicated that ABA is an important factor regulating the effects of GO on the growth and development of Brassica napus L., and that ETH and SA pathways may be potential pathways involved in the response of rape seedlings to GO treatment.

Metabolic prediction of important agronomic traits in hybrid rice (Oryza sativa L.).

Hybrid crops have contributed greatly to improvements in global food and fodder production over the past several decades. Nevertheless, the growing population and changing climate have produced food crises and energy shortages. Breeding new elite hybrid varieties is currently an urgent task, but present breeding procedures are time-consuming and labour-intensive. In this study, parental metabolic information was utilized to predict three polygenic traits in hybrid rice. A complete diallel cross population consisting of eighteen rice inbred lines was constructed, and the hybrids' plant height, heading date and grain yield per plant were predicted using 525 metabolites. Metabolic prediction models were built using the partial least square regression method, with predictive abilities ranging from 0.858 to 0.977 for the hybrid phenotypes, relative heterosis, and specific combining ability. Only slight changes in predictive ability were observed between hybrid populations, and nearly no changes were detected between reciprocal hybrids. The outcomes of prediction of the three highly polygenic traits demonstrated that metabolic prediction was an accurate (high predictive abilities) and efficient (unaffected by population genetic structures) strategy for screening promising superior hybrid rice. Exploitation of this pre-hybridization strategy may contribute to rice production improvement and accelerate breeding programs.

Fine Mapping of Two Additive Effect Genes for Awn Development in Rice (Oryza sativa L.).

Awns, important domestication and agronomic traits in rice (Oryza sativa L.), are conferred by polygenes and the environment. Near isogenic line (NIL) pairs BM33 and BM38 were constructed from crosses between awnless japonica cv Nipponbare as recurrent parent, and lines SLG or Funingxiaohongmang (awned japonica accessions), respectively, as donors. In order to study the genetic and molecular mechanism of awning, two unknown, independent genes with additive effects were identified in a cross between the NILs. To map and clone the two genes, a BC4F4 population of 8,103 individuals and a BC4F6 population of 11,206 individuals were constructed. Awn3-1 was fine mapped to a 101.13 kb genomic region between Indel marker In316 and SNP marker S9-1 on chromosome 3. Nine predicted genes in the interval were annotated in the Rice Annotation Project Database (RAP-DB), and Os03g0418600 was identified as the most likely candidate for Awn3-1 through sequence comparisons and RT-PCR assays. Awn4-2 was fine mapped to a 62.4 kb genomic region flanked by simple sequence repeat (SSR) marker M1126 and Indel maker In73 on chromosome 4L. This region contained the previously reported gene An-1 that regulates awn development. Thus, An-1 may be the candidate gene of Awn4-2. These results will facilitate cloning of the awn genes and thereby provide an understanding of the molecular basis of awn development.

Hierarchical additive effects on heterosis in rice (Oryza sativa L.).

Exploitation of heterosis in crops has contributed greatly to improvement in global food and energy production. In spite of the pervasive importance of heterosis, a complete understanding of its mechanisms has remained elusive. In this study, a small test-crossed rice population was constructed to investigate the formation mechanism of heterosis for 13 traits. The results of the relative mid-parent heterosis and modes of inheritance of all investigated traits demonstrated that additive effects were the foundation of heterosis for complex traits in a hierarchical structure, and multiplicative interactions among the component traits were the framework of heterosis in complex traits. Furthermore, new balances between unit traits and related component traits provided hybrids with the opportunity to achieve an optimal degree of heterosis for complex traits. This study dissected heterosis of both reproductive and vegetative traits from the perspective of hierarchical structure for the first time. Additive multiplicative interactions of component traits were proven to be the origin of heterosis in complex traits. Meanwhile, more attention should be paid to component traits, rather than complex traits, in the process of revealing the mechanism of heterosis.

Balance between a higher degree of heterosis and increased reproductive isolation: a strategic design for breeding inter-subspecific hybrid rice.

The application of heterosis (hybrid vigor) has brought great success to plant breeding, particularly of hybrid rice, achieving significant yield increases. Attempts to explore the heterosis of inter-subspecific hybrids between indica and japonica rice, which result in even greater yield increases, have greatly increased in the past decades. However, because of the reduced seed setting rate in F1 hybrids as a result of increased reproductive isolation, the application of inter-subspecific hybrids in rice has slowed. Understanding the balance between heterosis and the reproductive isolation of inter-subspecific hybrids will facilitate the strategic design of inter-subspecific hybrid breeding. In this study, five indica and seven japonica rice varieties were chosen as the parental lines of a complete diallel mating design. Data from six group traits from all of the hybrids and inbred lines were collected. We found that the grain weight per plant, grain number per panicle, tiller per plant, thousand grain weight and plant height, which reflected increased heterosis, were associated with the genetic divergence index (GDI) of the parents. Meanwhile, owing to the reduced seed setting rate, which was also associated with the parents' GDI, the grain production of the hybrids was negatively affected. After analyzing the relationships between the GDI of indica-japonica parents and the grain weight per plant of the F1 hybrids, an ideal GDI value (0.37) for the two indica-japonica parents that could provide an optimal balance between the inter-subspecific heterosis and reproductive isolation was proposed. Our findings will help in the strategic design of an inter-subspecific hybrid rice breeding program by identifying the ideal indica and japonica parents for a hybrid combination to achieve hybrid rice with an optimal yield. This strategic design of an inter-subspecific hybrid rice breeding program will be time saving and cost effective.

[Observation on short and long-term effects of cervical spondylotic radiculopathy treated with abdominal acupuncture plus Long's bone-setting manipulation].

OBJECTIVE: To compare the short and long-term therapeutic effects on cervical spondylotic radiculopathy (CSR) treated with simple Long's bone-setting manipulation, abdominal acupuncture and abdominal acupuncture plus Long's bone-setting manipulation. METHODS: One hundred and eighty cases of CSR were randomly allocated into abdominal acupuncture plus bone-setting group (combined therapy group), bone-setting group and abdominal acupuncture group, 60 cases in each group. In combined therapy group, the abdominal acupuncture and Long's bone-setting were applied in combination. Abdominal acupuncture was applied to Zhongwan (CV 12), Guanyuan (CV 4), Shiguan (KI 18), Shangqu (KI 17), etc. Long's manipulation, such as bone-setting in head-upward posture and bone-setting in head-lateral posture, was adopted. In bone-setting group and abdominal acupuncture group, Long's bone-setting manipulation and abdominal acupuncture were adopted simply and respectively. The clinical therapeutic effects were compared after 2 courses of treatment (short-term) and 1-month after treatment (long-term) among groups. RESULTS: The short and long-term curative and markedly effective rates in combined therapy group were 80.7% (46/57) and 68.4% (39/57) respectively, which were better than those of 63.64% (35/55), 30.9% (17/55) in bone-setting group and 58.9% (33/56), 50.0% (28/56) in abdominal acupuncture group, separately (all P < 0.05). Moreover, the long-term curative and markedly effective rate in abdominal acupuncture group was superior to that in bone-setting group (P < 0.05). CONCLUSION: Abdominal acupuncture plus Long's bone-setting manipulation has significant efficacy of either short or long-term on CSR, which is superior to the efficacy of either simple abdominal acupuncture or Long's bone-setting manipulation and indicates superimposed effect. Hence, it is one of the better approaches in CSR treatment.

[Clinical observation on acupuncture for treatment of hypertension of phlegm-stasis blocking collateral type].

OBJECTIVE: To explore the clinical therapeutic effect of acupuncture on hypertension of phlegm-stasis blocking collateral type and the mechanism. METHODS: Sixty cases of hypertension were randomly divided into a treatment group and a control group, 30 cases in each group. The treatment group were treated with acupuncture at Fengchi (GB 20), Quchi (LI 11), Neiguan (PC 6), Zusanli (ST 36), Fenglong (ST 40), Taicehong (LR 3), and oral administration of Captori, and the control group only with Captoril, for 4 therapeutic courses. The changes of blood pressure, clinical symptoms, hemorheologic parameters, C-reactive protein (CRP), TC, TG, HDL and LDL levels after treatment were observed. RESULTS: In the treatment group, blood pressure significantly decreased, and contents of CRP, TC, TG and LDHD decrease, the hemorheologic parameters improved and HDL level increased. Both the cumulative scores of clinical symptoms and the therapeutic effect for each symptom were improved significantly, with very significant differences as compared with those before treatment and the control group (P < 0.01). CONCLUSION: Acupuncture treatment has obvious effect of decreasing blood pressure, and reverses or delays the course of atherosclerosis, which mechanisms are possibly related with the function of protecting vascular endothelium.