Isochorismate synthase(ICS),a key rate-limiting enzyme in the salicylic acid(SA)biosynthesis pathway in plants,is essential for plant growth and defense against diseases.However,there has been no report on ICS in suga...Isochorismate synthase(ICS),a key rate-limiting enzyme in the salicylic acid(SA)biosynthesis pathway in plants,is essential for plant growth and defense against diseases.However,there has been no report on ICS in sugarcane(Saccharum spp.).In this study,18 SsICSs,42 ShICSs,and 36 SzICSs were identified from the genomes of sugarcane AP85-441(Saccharum spontaneum),XTT22(Saccharum spp.hybrid cultivar),and ZZ1(Saccharum spp.hybrid cultivar),respectively.These were phylogenetically divided into three groups,forming distinct clades that were evolutionarily divergent from those in dicotyledonous species.The evolutionary profile of the ICS gene family suggested expansion through whole-genome duplication/segmental events and strong purifying selection.Promoter cis-element and transcriptome analyses indicated that the ICS gene family responded to disease stress.We cloned the ScICS(isochorismate synthase)gene from sugarcane cultivar XTT22 leaves,and found it was localized in chloroplasts.In vivo and in vitro interaction studies revealed an interaction between ScICS and an ScMYB transcription factor.We showed that ScWRKY28 positively regulated ScICS expression by binding to its promoter.ScICS overexpression in transgenic tobacco confirmed its effectiveness in enhancing disease resistance.There was a significant increase in SA content following pathogen infection along with activation of downstream signaling pathways and defense mechanisms.This study establishes the groundwork for functional studies of sugarcane ICS genes and enhances our understanding of the mechanisms of disease resistance in sugarcane.展开更多
Sagittaria trifolia L.is a perennial aquatic herb that primarily reproduces clonally and through generative propagation.In recent years,S.trifolia has evolved a drastic resistance to acetohydroxy acid synthase(AHAS)-i...Sagittaria trifolia L.is a perennial aquatic herb that primarily reproduces clonally and through generative propagation.In recent years,S.trifolia has evolved a drastic resistance to acetohydroxy acid synthase(AHAS)-inhibiting herbicides in Northeast China.The phylogeographic patterns of S.trifolia with 31 purified resistance genotypes and five sensitive genotypes using chloroplast DNA(cpDNA)atpB-rbcL intergenic spacers were studied.Five haplotypes were characterized,and two of them were widely distributed in 36 genotypes.The dose response to bensulfuron-methyl showed that the GR50 ranged from 2.07 g a.i.·hm^(-2) to 220.15 g a.i.·hm^(-2).Sequencing of the AHAS gene indicated that 17 genotypes with the Pro197 mutation were distributed in haplotype 1,six genotypes with the Trp574 mutation were distributed in haplotype 3,and 13 genotypes with the wild AHAS gene were distributed in haplotypes 2,4 and 5.In the minimum-spanning network,the ancestral haplotypes 1 and 2 were widely distributed.Two primary clades were separated in the Bayes tree,and the result was consistent with the maximum likelihood tree.展开更多
In order to isolate the total cDNA of rice (Oryza sativa L.) epsps gene, RT-PCR was carried out with template of rice first-strand cDNA and primers designed according to rice EPSP synthase genomic sequence obtained in...In order to isolate the total cDNA of rice (Oryza sativa L.) epsps gene, RT-PCR was carried out with template of rice first-strand cDNA and primers designed according to rice EPSP synthase genomic sequence obtained in previous study. A 1 585-bp cDNA fragment was amplified and cloned. The 1 585-bp cDNA contains an open reading frame (ORF) comprising of 1 533 nucleotides (nt) which encodes a 511 residue polypepetides, including 67 amino acids chloroplast transit peptide and 444 amino acids EPSP synthase mature peptide. A comparison between the EPSP synthase of different sources indicates that the mature peptide shows more than 51% identity except for the fungi EPSP synthase and the transit peptide shows considerably less sequence conservation. The copy number of rice epsps gene is estimated to be one copy per haploid rice genome using southern blot. RT-PCR indicated that rice epsps gene is expressed in rice leaves, endosperms and roots and has the highest expression level in leaves.展开更多
A 1 886 bp full-length sesquiterpene synthase (AaSES) cDNA was cloned front a high-yield Artemisia annua L. strain 001 by a rapid amplification of cDNA end (RACE) strategy. AaSES is 59% identical to Artemisia cyclase ...A 1 886 bp full-length sesquiterpene synthase (AaSES) cDNA was cloned front a high-yield Artemisia annua L. strain 001 by a rapid amplification of cDNA end (RACE) strategy. AaSES is 59% identical to Artemisia cyclase cDNA clone cASC125, 50% identical to epi-cedrol synthase from A. annua , 48% identical to amorpha-4, 11-diene synthase from A. annua, 39% identical to the 5-epi-aristolechene synthase from tabacco, 38 % identical to vetispiradiene synthase front H. muticus, 41 % identical to the, delta-cadinene synthase from cotton. The coding region of the cDNA was inserted into a procaryotic expression vector pET-30a and overexpressed in E. coli BL21 ( DE3). The cyclase proteins extracted front bacterial culture were found largely in an insoluble protein fraction. AaSES expresses in leaves, stems a-rid flowers, not in roots as indicated by Northern blotting analysis.展开更多
Border-associated macrophages are located at the interface between the brain and the periphery, including the perivascular spaces, choroid plexus, and meninges. Until recently, the functions of border-associated macro...Border-associated macrophages are located at the interface between the brain and the periphery, including the perivascular spaces, choroid plexus, and meninges. Until recently, the functions of border-associated macrophages have been poorly understood and largely overlooked. However, a recent study reported that border-associated macrophages participate in stroke-induced inflammation, although many details and the underlying mechanisms remain unclear. In this study, we performed a comprehensive single-cell analysis of mouse border-associated macrophages using sequencing data obtained from the Gene Expression Omnibus(GEO) database(GSE174574 and GSE225948). Differentially expressed genes were identified, and enrichment analysis was performed to identify the transcription profile of border-associated macrophages. CellChat analysis was conducted to determine the cell communication network of border-associated macrophages. Transcription factors were predicted using the ‘pySCENIC' tool. We found that, in response to hypoxia, borderassociated macrophages underwent dynamic transcriptional changes and participated in the regulation of inflammatory-related pathways. Notably, the tumor necrosis factor pathway was activated by border-associated macrophages following ischemic stroke. The pySCENIC analysis indicated that the activity of signal transducer and activator of transcription 3(Stat3) was obviously upregulated in stroke, suggesting that Stat3 inhibition may be a promising strategy for treating border-associated macrophages-induced neuroinflammation. Finally, we constructed an animal model to investigate the effects of border-associated macrophages depletion following a stroke. Treatment with liposomes containing clodronate significantly reduced infarct volume in the animals and improved neurological scores compared with untreated animals. Taken together, our results demonstrate comprehensive changes in border-associated macrophages following a stroke, providing a theoretical basis for targeting border-associated macrophages-induced neuroinflammation in stroke treatment.展开更多
Background:Receptor-interacting protein kinases(RIPKs)regulate cell death,inflammation,and immune responses,yet their roles in cancer are not fully understood.This study investigates the expression,genomic alterations...Background:Receptor-interacting protein kinases(RIPKs)regulate cell death,inflammation,and immune responses,yet their roles in cancer are not fully understood.This study investigates the expression,genomic alterations,and functional implications of RIPK family members across various cancers.Methods:We collected multi-omics data from The Cancer Genome Atlas and other public databases,including gene expression,copy number variation(CNV),mutation,methylation,tumor mutation burden(TMB),and microsatellite instability(MSI).Differential expression and survival analyses were performed using DESeq2 and Cox proportional hazards models.CNV and mutation data were analyzed with GISTIC2 and Mutect2,and methylation data with the ChAMP package.Correlations with TMB and MSI were assessed using Pearson coefficients,and gene set enrichment analysis was conducted with the MSigDB Hallmark gene sets.Results:RIPK family members show significant differential expression in various cancers,with RIPK1 and RIPK4 frequently altered.Survival analysis reveals heterogeneous impacts on overall survival.CNV and mutation analyses identify high alteration frequencies for RIPK2 and RIPK7,affecting gene expression.RIPK1 and RIPK7 are hypermethylated in several cancers,inversely correlating with RIPK3 expression.RIPK1,RIPK2,RIPK5,RIPK6,and RIPK7 correlate positively with TMB,while RIPK3 shows negative correlations in some cancers.MSI analysis indicates associations with DNA mismatch repair.G ene set enrichment analysis highlights immune-related pathway enrichment for RIPK1,RIPK2,RIPK3,and RIPK6,and cell proliferation and DNA repair pathways for RIPK4 and RIPK5.RIPK family members showed heterogeneous alterations across cancers:for example,RIPK7 was mutated in up to~15%of u terine c orpus e ndometrial c arcinoma and l ung s quamous c ell c arcinoma cases,and RIPK1 and RIPK7 exhibited frequent promoter hypermethylation in multiple tumor types.Several genes displayed context-dependent associations with overall survival and with TMB/MSI.Conclusion:This pan-cancer analysis of the RIPK family reveals their diverse roles and potential as biomarkers and therapeutic targets.The findings emphasize the importance of RIPK genes in tumorigenesis and suggest context-dependent functions across cancer types.Further studies are needed to explore their mechanisms in cancer development and clinical applications.展开更多
The sulfation and decomposition process has proven effective in selectively extracting lithium from lepidolite.It is essential to clarify the thermochemical behavior and kinetic parameters of decomposition reactions.A...The sulfation and decomposition process has proven effective in selectively extracting lithium from lepidolite.It is essential to clarify the thermochemical behavior and kinetic parameters of decomposition reactions.Accordingly,comprehensive kinetic study by employing thermalgravimetric analysis at various heating rates was presented in this paper.Two main weight loss regions were observed during heating.The initial region corresponded to the dehydration of crystal water,whereas the subsequent region with overlapping peaks involved complex decomposition reactions.The overlapping peaks were separated into two individual reaction peaks and the activation energy of each peak was calculated using isoconversional kinetics methods.The activation energy of peak 1 exhibited a continual increase as the reaction conversion progressed,while that of peak 2 steadily decreased.The optimal kinetic models,identified as belonging to the random nucleation and subsequent growth category,provided valuable insights into the mechanism of the decomposition reactions.Furthermore,the adjustment factor was introduced to reconstruct the kinetic mechanism models,and the reconstructed models described the kinetic mechanism model more accurately for the decomposition reactions.This study enhanced the understanding of the thermochemical behavior and kinetic parameters of the lepidolite sulfation product decomposition reactions,further providing theoretical basis for promoting the selective extraction of lithium.展开更多
This paper investigates the reliability of internal marine combustion engines using an integrated approach that combines Fault Tree Analysis(FTA)and Bayesian Networks(BN).FTA provides a structured,top-down method for ...This paper investigates the reliability of internal marine combustion engines using an integrated approach that combines Fault Tree Analysis(FTA)and Bayesian Networks(BN).FTA provides a structured,top-down method for identifying critical failure modes and their root causes,while BN introduces flexibility in probabilistic reasoning,enabling dynamic updates based on new evidence.This dual methodology overcomes the limitations of static FTA models,offering a comprehensive framework for system reliability analysis.Critical failures,including External Leakage(ELU),Failure to Start(FTS),and Overheating(OHE),were identified as key risks.By incorporating redundancy into high-risk components such as pumps and batteries,the likelihood of these failures was significantly reduced.For instance,redundant pumps reduced the probability of ELU by 31.88%,while additional batteries decreased the occurrence of FTS by 36.45%.The results underscore the practical benefits of combining FTA and BN for enhancing system reliability,particularly in maritime applications where operational safety and efficiency are critical.This research provides valuable insights for maintenance planning and highlights the importance of redundancy in critical systems,especially as the industry transitions toward more autonomous vessels.展开更多
AIM:To explore the causal relationship between several possible behavioral factors and high myopia(HM)using multivariable Mendelian randomization(MVMR)approach and to find the mediators among them with mediation analy...AIM:To explore the causal relationship between several possible behavioral factors and high myopia(HM)using multivariable Mendelian randomization(MVMR)approach and to find the mediators among them with mediation analysis.METHODS:The causal effects of several behavioral factors,including screen time,education time,time spent outdoors,and physical activity,on the risk of HM using univariable Mendelian randomization(MR)and MVMR analyses were first assessed.Genome-wide association study summary statistics of serum metabolites were also used in mediation analysis to determine the extent to which serum metabolites mediate the effects of behavioral factors on HM.RESULTS:MR analyses indicated that both increased time spent outdoors and a higher frequency of moderate physical activity significantly reduced the risk of HM.Further MVMR analysis confirmed that moderate physical activity independently contributed to a lower risk of HM.Additionally,MR analyses identified 13 serum metabolites significantly associated with HM,of which 12 were lipids and one was an amino acid derivative.Mediation analysis revealed that six lipid metabolites mediated the protective effects of moderate physical activity on HM,with the highest mediation proportion observed for 1-(1-enyl-palmitoyl)-GPC(p-16:0;30.83%).CONCLUSION:This study suggests that in addition to outdoor time,moderate physical activity habits may have an independent protective effect against HM and pointed to lipid metabolites as priority targets for the prevention due to low physical activity.These results emphasize the importance of physical activity and metabolic health in HM and underscore the need for further study of these complex associations.展开更多
Sucrose synthases(SUS) are a family of enzymes that play pivotal roles in carbon partitioning, sink strength and plant development. A total of 11 SUS genes have been identified in the genome of Malus domestica(Md SUSs...Sucrose synthases(SUS) are a family of enzymes that play pivotal roles in carbon partitioning, sink strength and plant development. A total of 11 SUS genes have been identified in the genome of Malus domestica(Md SUSs), and phylogenetic analysis revealed that the Md SUS genes were divided into three groups, named as SUS I, SUS II and SUS III, respectively. The SUS I and SUS III groups included four homologs each, whereas the SUS II group contained three homologs. SUS genes in the same group showed similar structural characteristics, such as exon number, size and length distribution. After assessing four different tissues, Md SUS1 s and Md SUS2.1 showed the highest expression in fruit, whereas Md SUS2.2/2.3 and Md SUS3 s exhibit the highest expression in shoot tips. Most Md SUSs showed decreased expression during fruit development, similar to SUS enzyme activity, but both Md SUS2.1 and Md SUS1.4 displayed opposite expression profiles. These results suggest that different Md SUS genes might play distinct roles in the sink-source sugar cycle and sugar utilization in apple sink tissues.展开更多
AIM:To summarize publication trends in the field of strabismus over the past 30y and predict future research hotspots.METHODS:A total of 2915 English-language articles and reviews on strabismus,published between 1993 ...AIM:To summarize publication trends in the field of strabismus over the past 30y and predict future research hotspots.METHODS:A total of 2915 English-language articles and reviews on strabismus,published between 1993 and 2022,were retrieved from the Web of Science Core Collection.Bibliometric analyses were performed using VOSviewer and CiteSpace software to explore publication trends,as well as the contributions and collaborative networks of countries/regions,authors,institutions,and journals.RESULTS:The annual number of publications on strabismus showed a consistent upward trend.The United States(USA)maintained a leading position in this research field while Republic of Korea and China emerged as rapidly advancing contributors over the last decade.The University of California,Los Angeles ranked as the most productive institution,and Jonathan M.Holmes from USA was the most productive author.Journal of AAPOS was the leading journal with the most strabismus publications,whereas the two most highly cited articles were both published in Ophthalmology.Co-occurrence analysis identified pivotal keywords and burst terms,including intermittent exotropia(IXT),acute acquired comitant esotropia(AACE),functional magnetic resonance imaging(fMRI),and surgical treatment,which were confirmed as predominant and frontier topics.CONCLUSION:This study provides a comprehensive bibliometric analysis of strabismus research,revealing the evolution of research hotspots over the past 30y and outlining several cutting-edge directions for future investigation.展开更多
To our knowledge,few reports on Demodex studied at the molecular level are available at present.In this study our group,for the first time,cloned,sequenced and analyzed the chitin synthase(CHS) gene fragments of Demod...To our knowledge,few reports on Demodex studied at the molecular level are available at present.In this study our group,for the first time,cloned,sequenced and analyzed the chitin synthase(CHS) gene fragments of Demodex folliculorum,Demodex brevis,and Demodex canis(three isolates from each species) from Xi'an China,by designing specific primers based on the only partial sequence of the CHS gene of D.canis from Japan,retrieved from GenBank.Results show that amplification was successful only in three D.canis isolates and one D.brevis isolate out of the nine Demodex isolates.The obtained fragments were sequenced to be 339 bp for D.canis and 338 bp for D.brevis.The CHS gene sequence similarities between the three Xi'an D.canis isolates and one Japanese D.canis isolate ranged from 99.7% to 100.0%,and those between four D.canis isolates and one D.brevis isolate were 99.1%-99.4%.Phylogenetic trees based on maximum parsimony(MP) and maximum likelihood(ML) methods shared the same clusters,according with the traditional classification.Two open reading frames(ORFs) were identified in each CHS gene sequenced,and their corresponding amino acid sequences were located at the catalytic domain.The relatively conserved sequences could be deduced to be a CHS class A gene,which is associated with chitin synthesis in the integument of Demodex mites.展开更多
In the flavonoid biosynthesis pathway, Chalcone synthase (CHS) is involved in the formation of the pigment and has been shown to be a rate-limiting enzyme for the synthesis of flavonoids. In this study, a PCR approach...In the flavonoid biosynthesis pathway, Chalcone synthase (CHS) is involved in the formation of the pigment and has been shown to be a rate-limiting enzyme for the synthesis of flavonoids. In this study, a PCR approach was used to clone a Chalcone synthases cDNA from flower of sweet osmanthus “Chenghong Dangui” and it was designated as OfCHS (O. fragrans, CHS). The cDNA was 1383 bp long and a coding sequence (CDS) of 1173 bp encoding a polypeptide of 391 amino acids with an estimated molecular mass of 39.9 kDa. The theoretical isoelectric point was 6.23. Phylogenetic analysis demonstrated that OfCHS clustered with Olea europaea, Solenostemon scutellarioides, Perilla frutescens, Antirrhinum majus and Digitalis lanata. We also detected the expression of OfCHS in different tissues in “Dangui” and in two cultivars with varied coloration, “Zi Yingui” and “Chenghong Dangui” at different floral stages using quantitative real-time PCR. We observed that OfCHS transcript was higher in leaves than in petals in “Dangui”. The transcripts of OfCHS in “Zi Yingui” petals were higher than those in “Dangui” at three stages especially at xianyan stage and there was no significant difference between the two cultivars in the full flowering stage. “Chenghong Dangui” has a relatively high anthocyanin content compared to “Zi Yingui”. The relative amount of anthocyanin of “Chenghong Dangui” initially increases, and then decreases during the bloom period. However, the expression of CHS is the highest at the initial flowering stage. These data suggest that the OfCHS does not play a key role in the accumulation of total flavonoid in this cultivar. These data could contribute to explain the different accumulation of flavonoids in petals of the two cultivars.展开更多
Sucrose phosphate synthase(SPS)is a rate-limiting enzyme that works in conjunction with sucrose-6-phosphate phosphatase(SPP)for sucrose synthesis,and it plays an essential role in energy provisioning during growth and...Sucrose phosphate synthase(SPS)is a rate-limiting enzyme that works in conjunction with sucrose-6-phosphate phosphatase(SPP)for sucrose synthesis,and it plays an essential role in energy provisioning during growth and development in plants as well as improving fruit quality.However,studies on the systematic analysis and evolutionary pattern of the SPS gene family in apple are still lacking.In the present study,a total of seven MdSPS and four MdSPP genes were identified from the Malus domestica genome GDDH13 v1.1.The gene structures and their promoter cis-elements,protein conserved motifs,subcellular localizations,physiological functions and biochemical properties were analyzed.A chromosomal location and gene-duplication analysis demonstrated that whole-genome duplication(WGD)and segmental duplication played vital roles in MdSPS gene family expansion.The Ka/Ks ratio of pairwise MdSPS genes indicated that the members of this family have undergone strong purifying selection during domestication.Furthermore,three SPS gene subfamilies were classified based on phylogenetic relationships,and old gene duplications and significantly divergent evolutionary rates were observed among the SPS gene subfamilies.In addition,a major gene related to sucrose accumulation(MdSPSA2.3)was identified according to the highly consistent trends in the changes of its expression in four apple varieties(‘Golden Delicious’,‘Fuji’,‘Qinguan’and‘Honeycrisp’)and the correlation between gene expression and soluble sugar content during fruit development.Furthermore,the virus-induced silencing of MdSPSA2.3 confirmed its function in sucrose accumulation in apple fruit.The present study lays a theoretical foundation for better clarifying the biological functions of the MdSPS genes during apple fruit development.展开更多
Objective:To clone,express and purify 2—methylcitrale synthase(Rv1131) gene of Mycobacterium tuberculosis(M.tuberculosis) and to study its structural characteristics using various bioinformatics tools.Methods:Rv1131 ...Objective:To clone,express and purify 2—methylcitrale synthase(Rv1131) gene of Mycobacterium tuberculosis(M.tuberculosis) and to study its structural characteristics using various bioinformatics tools.Methods:Rv1131 gene was amplified In polymerase chain reaction using M.tuberculosis H37 Rv genomic DNA and cloned into pGEM-T easy vector and sequenced.The gene was sub-cloned in pET28 c vector,expressed in Escherichia coli RL22(E.coli BL21)(DE3) cells and the recombinant protein was identified by Western blotting.The protein was purified using Nickel affinity chromatography and the structural characteristics like sub-cellular localization.presence of transmembrane helices and secondary structure of the protein were predicted by bioinformaties tools.Tertiary structure of the protein and phylogenetic analysis was aiso established by in silico analysis.Results:The expression of the recombinant protein(Rv1131) was confirmed by western blotting using anti-HIS antibodies and the protein was purified from the soluble fraction.In silico analysis showed that the protein contains no signal peptide and transmembrane helices,Active site prediction showed that the protein has histidine and aspartie acid residues at 242.281 & 332 positions respectively.Phylogenetic analysis showed100%homology with major mycobacterial species.Secondary structure predicts 2-methyleitrate synthase contain 51.9%;alpha-helk.8.7%extended strand and 39.4%random coils.Tertiary structure of the protein was also established.Conclusions:The enzyme 2-methylcitrate synthase from M.tuberculosis H37 Rv has been successfully expressed and purified.The purified protein will further be utilized to develop assay methods lor screening new inhibitors.展开更多
Syringa species are important ornamentals with strong floral scent,of which monoterpenes are the main component.In this study,a new monoterpene synthase gene,named SoLIM,was collected from the flowers of Syringa oblat...Syringa species are important ornamentals with strong floral scent,of which monoterpenes are the main component.In this study,a new monoterpene synthase gene,named SoLIM,was collected from the flowers of Syringa oblata and S.oblata var.alba using a homologous cloning method.The full-length cDNA of SoLIM was1746 bp and encoded 581 amino acids.Sequence analysis showed that SoLIM contained the DDxxD and RRx8 W motifs,which are two typical conserved monoterpene synthase motifs,and was thus classified as belonging to the Tpsb subfamily.Using quantitative reverse-transcription PCR,SoLIM was significantly expressed in the petals and pistils of S.oblata and S.oblata var.alba,respectively.SoLIM expression peaked earlier than the D-limonene emissions in the diurnal experiments,but occurred later when D-limonene had peaked during the flowering phase,indicating that differences in SoLIM gene expression and D-limonene emissions existed.The synthesis of floral scent is thus associated with diverse regulatory mechanisms that require further investigation.展开更多
With the increasing promotion of simplified rapeseed cultivation in recent years,the development of cultivars with high resistance to herbicides is urgently needed.We previously developed M342,which shows sulfonylurea...With the increasing promotion of simplified rapeseed cultivation in recent years,the development of cultivars with high resistance to herbicides is urgently needed.We previously developed M342,which shows sulfonylurea herbicide resistance,by targeting acetohydroxyacid synthase(AHAS),a key enzyme in branched-chain amino acid synthesis.In the present study,we used a progeny line derived from M342 for an additional round of ethyl methane sulfonate mutagenesis,yielding the novel mutant DS3,which harbored two mutations in AHAS genes and showed high sulfonylurea resistance.One mutation was the substitution Trp574 Leu,as in M342,according to Arabidopsis protein sequencing.The other site was a newly recognized substitution,Pro197 Leu.A KASP marker targeting Pro197 Leu was developed and reliably predicted the response to sulfonylurea herbicides in the F2 population.The combination of Trp574 Leu and Pro197 Leu in DS3 produced a synergistic effect that greatly increased herbicide resistance.Analysis of the protein structures of AHAS1 and AHAS3 in wild-type and single-gene mutant plants revealed three-dimensional protein conformational changes that could account for differences in herbicide resistance characteristics including toxicity tolerance,AHAS enzyme activity,and AHAS gene expression.展开更多
Chalcone synthases （CHS, EC 2.3.1.74） are key enzymes that catalyze the first committed step in flavonoid biosynthesis. In this study, we isolated a chalcone synthase, named NtCHS6, from Nicotiana tabacum. This synt...Chalcone synthases （CHS, EC 2.3.1.74） are key enzymes that catalyze the first committed step in flavonoid biosynthesis. In this study, we isolated a chalcone synthase, named NtCHS6, from Nicotiana tabacum. This synthase shared high homology with the NSCHSL （Y14507） gene and contained most of the conserved active sites that are in CHS proteins. The phylogenetic analysis suggested that NtCHS6 protein shared a large genetic distance with other Solanaceae CHS proteins and was the most closely-related to an uncharacterized CHS from Solanum lycopersicum. The expression analysis indicated that NtCHS6 was abundantly expressed in leaves, especially in mature leaves. By scrutinizing its upstream promoter sequences, multiple cis-regulatory elements involved in light and drought responsive were detected. Furthermore, NtCHS6 expression decreased significantly under dark treatment and increased significantly under drought stress suggested that NtCHS6 expression exhibited both light responsiveness and drought responsiveness, and important roles in ultraviolet protection and drought tolerance. Our results might play展开更多
Isochorismate synthase(ICS) is a crucial enzyme in the salicylic acid(SA) synthesis pathway. The full-length complementary DNA(cDNA) sequence of the ICS gene was isolated from Artemisia annua L. The gene, named AaICS...Isochorismate synthase(ICS) is a crucial enzyme in the salicylic acid(SA) synthesis pathway. The full-length complementary DNA(cDNA) sequence of the ICS gene was isolated from Artemisia annua L. The gene, named AaICS1, contained a 1710-bp open reading frame, which encoded a protein with 570 amino acids. Bioinformatics and comparative study revealed that the polypeptide protein of AaICS1 had high homology with ICSs from other plant species. Southern blot analysis suggested that AaICS1 might be a single-copy gene. Analysis of the 1470-bp promoter of AaICS1 identified distinct cis-acting regulatory elements, including TC-rich repeats, MYB binding site(MBS), and TCA-elements. An analysis of AaICS1 transcript levels in multifarious tissues of A. annua using quantitative real-time polymerase chain reaction(qRT-PCR) showed that old leaves had the highest transcription levels. AaICS1 was up-regulated under wounding, drought, salinity, and SA treatments. This was corroborated by the presence of the predicted cis-acting elements in the promoter region of AaICS1. Overexpressing transgenic plants and RNA interference transgenic lines of AaICS1 were generated and their expression was compared. High-performance liquid chromatography(HPLC) results from leaf tissue of transgenic A. annua showed an increase in artemisinin content in the overexpressing plants. These results confirm that AaICS1 is involved in the isochorismate pathway.展开更多
The study aims to reveal phylogenetic and evolutionary relationship between aerobic anoxygenic phototrophic bacteria（AAnPB） and their relatives,anaerobic anoxygenic phototrophic bacteria（AnAnPB） and nonphototrophi...The study aims to reveal phylogenetic and evolutionary relationship between aerobic anoxygenic phototrophic bacteria（AAnPB） and their relatives,anaerobic anoxygenic phototrophic bacteria（AnAnPB） and nonphototrophic bacteria（NPB,which had high homology of 16S rDNA gene with AAnPB and fell into the same genus）,and validate reliability and usefulness of farnesyl pyrophosphate synthase（FPPS） gene for the phylogenetic determination.FPPS genes with our modified primers and 16S rDNA genes with general primers,were amplified and sequenced or retrieved from GenBank database.In contrast to 16S rDNA gene phylogenetic tree,AAnPB were grouped into two clusters and one branch alone with no intermingling with NPB and AnAnPB in the tree constructed on FPPS.One branch of AAnPB,in both trees,was located closer to outgroup species than AnAnPB,which implicated that some AAnPB would be diverged earlier in FPPS evolutionary history than AnAnPB and NPB.Some AAnPB and NPB were closer located in both trees and this suggested that they were the closer relatives than AnAnPB.Combination codon usage in FPPS with phylogenetic analysis,the results indicates that FPPS gene and 16S rRNA gene have similar evolutionary pattern but the former seems to be more reliable and useful in determining the phylogenic and evolutionary relationship between AAnPB and their relatives.This is the first attempt to use a molecular marker beside 16S rRNA gene for studying the phylogeny of AAnPB,and the study may also be helpful in understanding the evolutionary relationship among phototrophic microbes and the trends of photosynthetic genes transfer.展开更多
基金supported by the National Natural Science Foundation of China(32472166,32172063,and 31771861)the Innovative Team Construction Project of the Modern Agricultural Industry Technology System in Guangdong Province by Agricultural Product Units(Sugarcane and Sisal Industry Technology System,2024CXTD03-06)South China Agricultural University Students Innovation and Entrepreneurship Training Program(2024105641195)。
文摘Isochorismate synthase(ICS),a key rate-limiting enzyme in the salicylic acid(SA)biosynthesis pathway in plants,is essential for plant growth and defense against diseases.However,there has been no report on ICS in sugarcane(Saccharum spp.).In this study,18 SsICSs,42 ShICSs,and 36 SzICSs were identified from the genomes of sugarcane AP85-441(Saccharum spontaneum),XTT22(Saccharum spp.hybrid cultivar),and ZZ1(Saccharum spp.hybrid cultivar),respectively.These were phylogenetically divided into three groups,forming distinct clades that were evolutionarily divergent from those in dicotyledonous species.The evolutionary profile of the ICS gene family suggested expansion through whole-genome duplication/segmental events and strong purifying selection.Promoter cis-element and transcriptome analyses indicated that the ICS gene family responded to disease stress.We cloned the ScICS(isochorismate synthase)gene from sugarcane cultivar XTT22 leaves,and found it was localized in chloroplasts.In vivo and in vitro interaction studies revealed an interaction between ScICS and an ScMYB transcription factor.We showed that ScWRKY28 positively regulated ScICS expression by binding to its promoter.ScICS overexpression in transgenic tobacco confirmed its effectiveness in enhancing disease resistance.There was a significant increase in SA content following pathogen infection along with activation of downstream signaling pathways and defense mechanisms.This study establishes the groundwork for functional studies of sugarcane ICS genes and enhances our understanding of the mechanisms of disease resistance in sugarcane.
基金Supported by the"Young Talents"Project of Northeast Agricultural University(22QC04)the Domestic Post Training Excellent Program of Northeast Agricultural University(23ZYZZ0706)。
文摘Sagittaria trifolia L.is a perennial aquatic herb that primarily reproduces clonally and through generative propagation.In recent years,S.trifolia has evolved a drastic resistance to acetohydroxy acid synthase(AHAS)-inhibiting herbicides in Northeast China.The phylogeographic patterns of S.trifolia with 31 purified resistance genotypes and five sensitive genotypes using chloroplast DNA(cpDNA)atpB-rbcL intergenic spacers were studied.Five haplotypes were characterized,and two of them were widely distributed in 36 genotypes.The dose response to bensulfuron-methyl showed that the GR50 ranged from 2.07 g a.i.·hm^(-2) to 220.15 g a.i.·hm^(-2).Sequencing of the AHAS gene indicated that 17 genotypes with the Pro197 mutation were distributed in haplotype 1,six genotypes with the Trp574 mutation were distributed in haplotype 3,and 13 genotypes with the wild AHAS gene were distributed in haplotypes 2,4 and 5.In the minimum-spanning network,the ancestral haplotypes 1 and 2 were widely distributed.Two primary clades were separated in the Bayes tree,and the result was consistent with the maximum likelihood tree.
文摘In order to isolate the total cDNA of rice (Oryza sativa L.) epsps gene, RT-PCR was carried out with template of rice first-strand cDNA and primers designed according to rice EPSP synthase genomic sequence obtained in previous study. A 1 585-bp cDNA fragment was amplified and cloned. The 1 585-bp cDNA contains an open reading frame (ORF) comprising of 1 533 nucleotides (nt) which encodes a 511 residue polypepetides, including 67 amino acids chloroplast transit peptide and 444 amino acids EPSP synthase mature peptide. A comparison between the EPSP synthase of different sources indicates that the mature peptide shows more than 51% identity except for the fungi EPSP synthase and the transit peptide shows considerably less sequence conservation. The copy number of rice epsps gene is estimated to be one copy per haploid rice genome using southern blot. RT-PCR indicated that rice epsps gene is expressed in rice leaves, endosperms and roots and has the highest expression level in leaves.
文摘A 1 886 bp full-length sesquiterpene synthase (AaSES) cDNA was cloned front a high-yield Artemisia annua L. strain 001 by a rapid amplification of cDNA end (RACE) strategy. AaSES is 59% identical to Artemisia cyclase cDNA clone cASC125, 50% identical to epi-cedrol synthase from A. annua , 48% identical to amorpha-4, 11-diene synthase from A. annua, 39% identical to the 5-epi-aristolechene synthase from tabacco, 38 % identical to vetispiradiene synthase front H. muticus, 41 % identical to the, delta-cadinene synthase from cotton. The coding region of the cDNA was inserted into a procaryotic expression vector pET-30a and overexpressed in E. coli BL21 ( DE3). The cyclase proteins extracted front bacterial culture were found largely in an insoluble protein fraction. AaSES expresses in leaves, stems a-rid flowers, not in roots as indicated by Northern blotting analysis.
基金supported by Qingdao Key Medical and Health Discipline ProjectThe Intramural Research Program of the Affiliated Hospital of Qingdao University,No. 4910Qingdao West Coast New Area Science and Technology Project,No. 2020-55 (all to SW)。
文摘Border-associated macrophages are located at the interface between the brain and the periphery, including the perivascular spaces, choroid plexus, and meninges. Until recently, the functions of border-associated macrophages have been poorly understood and largely overlooked. However, a recent study reported that border-associated macrophages participate in stroke-induced inflammation, although many details and the underlying mechanisms remain unclear. In this study, we performed a comprehensive single-cell analysis of mouse border-associated macrophages using sequencing data obtained from the Gene Expression Omnibus(GEO) database(GSE174574 and GSE225948). Differentially expressed genes were identified, and enrichment analysis was performed to identify the transcription profile of border-associated macrophages. CellChat analysis was conducted to determine the cell communication network of border-associated macrophages. Transcription factors were predicted using the ‘pySCENIC' tool. We found that, in response to hypoxia, borderassociated macrophages underwent dynamic transcriptional changes and participated in the regulation of inflammatory-related pathways. Notably, the tumor necrosis factor pathway was activated by border-associated macrophages following ischemic stroke. The pySCENIC analysis indicated that the activity of signal transducer and activator of transcription 3(Stat3) was obviously upregulated in stroke, suggesting that Stat3 inhibition may be a promising strategy for treating border-associated macrophages-induced neuroinflammation. Finally, we constructed an animal model to investigate the effects of border-associated macrophages depletion following a stroke. Treatment with liposomes containing clodronate significantly reduced infarct volume in the animals and improved neurological scores compared with untreated animals. Taken together, our results demonstrate comprehensive changes in border-associated macrophages following a stroke, providing a theoretical basis for targeting border-associated macrophages-induced neuroinflammation in stroke treatment.
基金supported by grants from the Tianjin Health Technology Project(Grant no.2022QN106).
文摘Background:Receptor-interacting protein kinases(RIPKs)regulate cell death,inflammation,and immune responses,yet their roles in cancer are not fully understood.This study investigates the expression,genomic alterations,and functional implications of RIPK family members across various cancers.Methods:We collected multi-omics data from The Cancer Genome Atlas and other public databases,including gene expression,copy number variation(CNV),mutation,methylation,tumor mutation burden(TMB),and microsatellite instability(MSI).Differential expression and survival analyses were performed using DESeq2 and Cox proportional hazards models.CNV and mutation data were analyzed with GISTIC2 and Mutect2,and methylation data with the ChAMP package.Correlations with TMB and MSI were assessed using Pearson coefficients,and gene set enrichment analysis was conducted with the MSigDB Hallmark gene sets.Results:RIPK family members show significant differential expression in various cancers,with RIPK1 and RIPK4 frequently altered.Survival analysis reveals heterogeneous impacts on overall survival.CNV and mutation analyses identify high alteration frequencies for RIPK2 and RIPK7,affecting gene expression.RIPK1 and RIPK7 are hypermethylated in several cancers,inversely correlating with RIPK3 expression.RIPK1,RIPK2,RIPK5,RIPK6,and RIPK7 correlate positively with TMB,while RIPK3 shows negative correlations in some cancers.MSI analysis indicates associations with DNA mismatch repair.G ene set enrichment analysis highlights immune-related pathway enrichment for RIPK1,RIPK2,RIPK3,and RIPK6,and cell proliferation and DNA repair pathways for RIPK4 and RIPK5.RIPK family members showed heterogeneous alterations across cancers:for example,RIPK7 was mutated in up to~15%of u terine c orpus e ndometrial c arcinoma and l ung s quamous c ell c arcinoma cases,and RIPK1 and RIPK7 exhibited frequent promoter hypermethylation in multiple tumor types.Several genes displayed context-dependent associations with overall survival and with TMB/MSI.Conclusion:This pan-cancer analysis of the RIPK family reveals their diverse roles and potential as biomarkers and therapeutic targets.The findings emphasize the importance of RIPK genes in tumorigenesis and suggest context-dependent functions across cancer types.Further studies are needed to explore their mechanisms in cancer development and clinical applications.
基金financially supported by the National Natural Science Foundation of China(Nos.52034002 and U2202254)the Fundamental Research Funds for the Central Universities,China(No.FRF-TT-19-001)。
文摘The sulfation and decomposition process has proven effective in selectively extracting lithium from lepidolite.It is essential to clarify the thermochemical behavior and kinetic parameters of decomposition reactions.Accordingly,comprehensive kinetic study by employing thermalgravimetric analysis at various heating rates was presented in this paper.Two main weight loss regions were observed during heating.The initial region corresponded to the dehydration of crystal water,whereas the subsequent region with overlapping peaks involved complex decomposition reactions.The overlapping peaks were separated into two individual reaction peaks and the activation energy of each peak was calculated using isoconversional kinetics methods.The activation energy of peak 1 exhibited a continual increase as the reaction conversion progressed,while that of peak 2 steadily decreased.The optimal kinetic models,identified as belonging to the random nucleation and subsequent growth category,provided valuable insights into the mechanism of the decomposition reactions.Furthermore,the adjustment factor was introduced to reconstruct the kinetic mechanism models,and the reconstructed models described the kinetic mechanism model more accurately for the decomposition reactions.This study enhanced the understanding of the thermochemical behavior and kinetic parameters of the lepidolite sulfation product decomposition reactions,further providing theoretical basis for promoting the selective extraction of lithium.
基金supported by Istanbul Technical University(Project No.45698)supported through the“Young Researchers’Career Development Project-training of doctoral students”of the Croatian Science Foundation.
文摘This paper investigates the reliability of internal marine combustion engines using an integrated approach that combines Fault Tree Analysis(FTA)and Bayesian Networks(BN).FTA provides a structured,top-down method for identifying critical failure modes and their root causes,while BN introduces flexibility in probabilistic reasoning,enabling dynamic updates based on new evidence.This dual methodology overcomes the limitations of static FTA models,offering a comprehensive framework for system reliability analysis.Critical failures,including External Leakage(ELU),Failure to Start(FTS),and Overheating(OHE),were identified as key risks.By incorporating redundancy into high-risk components such as pumps and batteries,the likelihood of these failures was significantly reduced.For instance,redundant pumps reduced the probability of ELU by 31.88%,while additional batteries decreased the occurrence of FTS by 36.45%.The results underscore the practical benefits of combining FTA and BN for enhancing system reliability,particularly in maritime applications where operational safety and efficiency are critical.This research provides valuable insights for maintenance planning and highlights the importance of redundancy in critical systems,especially as the industry transitions toward more autonomous vessels.
基金Supported by the Central High Level Hospital Clinical Research Funding(No.BJ-2024-089).
文摘AIM:To explore the causal relationship between several possible behavioral factors and high myopia(HM)using multivariable Mendelian randomization(MVMR)approach and to find the mediators among them with mediation analysis.METHODS:The causal effects of several behavioral factors,including screen time,education time,time spent outdoors,and physical activity,on the risk of HM using univariable Mendelian randomization(MR)and MVMR analyses were first assessed.Genome-wide association study summary statistics of serum metabolites were also used in mediation analysis to determine the extent to which serum metabolites mediate the effects of behavioral factors on HM.RESULTS:MR analyses indicated that both increased time spent outdoors and a higher frequency of moderate physical activity significantly reduced the risk of HM.Further MVMR analysis confirmed that moderate physical activity independently contributed to a lower risk of HM.Additionally,MR analyses identified 13 serum metabolites significantly associated with HM,of which 12 were lipids and one was an amino acid derivative.Mediation analysis revealed that six lipid metabolites mediated the protective effects of moderate physical activity on HM,with the highest mediation proportion observed for 1-(1-enyl-palmitoyl)-GPC(p-16:0;30.83%).CONCLUSION:This study suggests that in addition to outdoor time,moderate physical activity habits may have an independent protective effect against HM and pointed to lipid metabolites as priority targets for the prevention due to low physical activity.These results emphasize the importance of physical activity and metabolic health in HM and underscore the need for further study of these complex associations.
基金supported in part by the National Natural Science Foundation of China (31372038)the Natural Basic Research Plan in Shaanxi Province of China (2015JQ3082)
文摘Sucrose synthases(SUS) are a family of enzymes that play pivotal roles in carbon partitioning, sink strength and plant development. A total of 11 SUS genes have been identified in the genome of Malus domestica(Md SUSs), and phylogenetic analysis revealed that the Md SUS genes were divided into three groups, named as SUS I, SUS II and SUS III, respectively. The SUS I and SUS III groups included four homologs each, whereas the SUS II group contained three homologs. SUS genes in the same group showed similar structural characteristics, such as exon number, size and length distribution. After assessing four different tissues, Md SUS1 s and Md SUS2.1 showed the highest expression in fruit, whereas Md SUS2.2/2.3 and Md SUS3 s exhibit the highest expression in shoot tips. Most Md SUSs showed decreased expression during fruit development, similar to SUS enzyme activity, but both Md SUS2.1 and Md SUS1.4 displayed opposite expression profiles. These results suggest that different Md SUS genes might play distinct roles in the sink-source sugar cycle and sugar utilization in apple sink tissues.
基金Supported by National Natural Science Foundation of China(No.82020108006,No.81730025).
文摘AIM:To summarize publication trends in the field of strabismus over the past 30y and predict future research hotspots.METHODS:A total of 2915 English-language articles and reviews on strabismus,published between 1993 and 2022,were retrieved from the Web of Science Core Collection.Bibliometric analyses were performed using VOSviewer and CiteSpace software to explore publication trends,as well as the contributions and collaborative networks of countries/regions,authors,institutions,and journals.RESULTS:The annual number of publications on strabismus showed a consistent upward trend.The United States(USA)maintained a leading position in this research field while Republic of Korea and China emerged as rapidly advancing contributors over the last decade.The University of California,Los Angeles ranked as the most productive institution,and Jonathan M.Holmes from USA was the most productive author.Journal of AAPOS was the leading journal with the most strabismus publications,whereas the two most highly cited articles were both published in Ophthalmology.Co-occurrence analysis identified pivotal keywords and burst terms,including intermittent exotropia(IXT),acute acquired comitant esotropia(AACE),functional magnetic resonance imaging(fMRI),and surgical treatment,which were confirmed as predominant and frontier topics.CONCLUSION:This study provides a comprehensive bibliometric analysis of strabismus research,revealing the evolution of research hotspots over the past 30y and outlining several cutting-edge directions for future investigation.
基金Project supported by the National Natural Science Foundation of China (No. 81271856)the National College of Innovative Experimental Projects of China (No. 101069819)
文摘To our knowledge,few reports on Demodex studied at the molecular level are available at present.In this study our group,for the first time,cloned,sequenced and analyzed the chitin synthase(CHS) gene fragments of Demodex folliculorum,Demodex brevis,and Demodex canis(three isolates from each species) from Xi'an China,by designing specific primers based on the only partial sequence of the CHS gene of D.canis from Japan,retrieved from GenBank.Results show that amplification was successful only in three D.canis isolates and one D.brevis isolate out of the nine Demodex isolates.The obtained fragments were sequenced to be 339 bp for D.canis and 338 bp for D.brevis.The CHS gene sequence similarities between the three Xi'an D.canis isolates and one Japanese D.canis isolate ranged from 99.7% to 100.0%,and those between four D.canis isolates and one D.brevis isolate were 99.1%-99.4%.Phylogenetic trees based on maximum parsimony(MP) and maximum likelihood(ML) methods shared the same clusters,according with the traditional classification.Two open reading frames(ORFs) were identified in each CHS gene sequenced,and their corresponding amino acid sequences were located at the catalytic domain.The relatively conserved sequences could be deduced to be a CHS class A gene,which is associated with chitin synthesis in the integument of Demodex mites.
文摘In the flavonoid biosynthesis pathway, Chalcone synthase (CHS) is involved in the formation of the pigment and has been shown to be a rate-limiting enzyme for the synthesis of flavonoids. In this study, a PCR approach was used to clone a Chalcone synthases cDNA from flower of sweet osmanthus “Chenghong Dangui” and it was designated as OfCHS (O. fragrans, CHS). The cDNA was 1383 bp long and a coding sequence (CDS) of 1173 bp encoding a polypeptide of 391 amino acids with an estimated molecular mass of 39.9 kDa. The theoretical isoelectric point was 6.23. Phylogenetic analysis demonstrated that OfCHS clustered with Olea europaea, Solenostemon scutellarioides, Perilla frutescens, Antirrhinum majus and Digitalis lanata. We also detected the expression of OfCHS in different tissues in “Dangui” and in two cultivars with varied coloration, “Zi Yingui” and “Chenghong Dangui” at different floral stages using quantitative real-time PCR. We observed that OfCHS transcript was higher in leaves than in petals in “Dangui”. The transcripts of OfCHS in “Zi Yingui” petals were higher than those in “Dangui” at three stages especially at xianyan stage and there was no significant difference between the two cultivars in the full flowering stage. “Chenghong Dangui” has a relatively high anthocyanin content compared to “Zi Yingui”. The relative amount of anthocyanin of “Chenghong Dangui” initially increases, and then decreases during the bloom period. However, the expression of CHS is the highest at the initial flowering stage. These data suggest that the OfCHS does not play a key role in the accumulation of total flavonoid in this cultivar. These data could contribute to explain the different accumulation of flavonoids in petals of the two cultivars.
基金supported by the National Natural Science Foundation of China (32172521)the Excellent Youth Science Foundation of Heilongjiang Province,China (YQ2023C006)+1 种基金the Talent Introduction Program of Northeast Agricultural University of Chinathe Collaborative Innovation System of the Agricultural Bio-economy in Heilongjiang Province,China
文摘Sucrose phosphate synthase(SPS)is a rate-limiting enzyme that works in conjunction with sucrose-6-phosphate phosphatase(SPP)for sucrose synthesis,and it plays an essential role in energy provisioning during growth and development in plants as well as improving fruit quality.However,studies on the systematic analysis and evolutionary pattern of the SPS gene family in apple are still lacking.In the present study,a total of seven MdSPS and four MdSPP genes were identified from the Malus domestica genome GDDH13 v1.1.The gene structures and their promoter cis-elements,protein conserved motifs,subcellular localizations,physiological functions and biochemical properties were analyzed.A chromosomal location and gene-duplication analysis demonstrated that whole-genome duplication(WGD)and segmental duplication played vital roles in MdSPS gene family expansion.The Ka/Ks ratio of pairwise MdSPS genes indicated that the members of this family have undergone strong purifying selection during domestication.Furthermore,three SPS gene subfamilies were classified based on phylogenetic relationships,and old gene duplications and significantly divergent evolutionary rates were observed among the SPS gene subfamilies.In addition,a major gene related to sucrose accumulation(MdSPSA2.3)was identified according to the highly consistent trends in the changes of its expression in four apple varieties(‘Golden Delicious’,‘Fuji’,‘Qinguan’and‘Honeycrisp’)and the correlation between gene expression and soluble sugar content during fruit development.Furthermore,the virus-induced silencing of MdSPSA2.3 confirmed its function in sucrose accumulation in apple fruit.The present study lays a theoretical foundation for better clarifying the biological functions of the MdSPS genes during apple fruit development.
基金supported by funding from Council of Scicntific and Industrial Researd(CSIR),India
文摘Objective:To clone,express and purify 2—methylcitrale synthase(Rv1131) gene of Mycobacterium tuberculosis(M.tuberculosis) and to study its structural characteristics using various bioinformatics tools.Methods:Rv1131 gene was amplified In polymerase chain reaction using M.tuberculosis H37 Rv genomic DNA and cloned into pGEM-T easy vector and sequenced.The gene was sub-cloned in pET28 c vector,expressed in Escherichia coli RL22(E.coli BL21)(DE3) cells and the recombinant protein was identified by Western blotting.The protein was purified using Nickel affinity chromatography and the structural characteristics like sub-cellular localization.presence of transmembrane helices and secondary structure of the protein were predicted by bioinformaties tools.Tertiary structure of the protein and phylogenetic analysis was aiso established by in silico analysis.Results:The expression of the recombinant protein(Rv1131) was confirmed by western blotting using anti-HIS antibodies and the protein was purified from the soluble fraction.In silico analysis showed that the protein contains no signal peptide and transmembrane helices,Active site prediction showed that the protein has histidine and aspartie acid residues at 242.281 & 332 positions respectively.Phylogenetic analysis showed100%homology with major mycobacterial species.Secondary structure predicts 2-methyleitrate synthase contain 51.9%;alpha-helk.8.7%extended strand and 39.4%random coils.Tertiary structure of the protein was also established.Conclusions:The enzyme 2-methylcitrate synthase from M.tuberculosis H37 Rv has been successfully expressed and purified.The purified protein will further be utilized to develop assay methods lor screening new inhibitors.
基金supported by the Project of Construction of Innovative Teams and Teacher Career Development for Universities and Colleges under Beijing Municipality(IDHT20180509)the National Natural Science foundation of China(31201645)+1 种基金the Beijing Municipal Natural Science Foundation(6172006)key project of Beijing Municipal Education Commission(KZ201510020021)
文摘Syringa species are important ornamentals with strong floral scent,of which monoterpenes are the main component.In this study,a new monoterpene synthase gene,named SoLIM,was collected from the flowers of Syringa oblata and S.oblata var.alba using a homologous cloning method.The full-length cDNA of SoLIM was1746 bp and encoded 581 amino acids.Sequence analysis showed that SoLIM contained the DDxxD and RRx8 W motifs,which are two typical conserved monoterpene synthase motifs,and was thus classified as belonging to the Tpsb subfamily.Using quantitative reverse-transcription PCR,SoLIM was significantly expressed in the petals and pistils of S.oblata and S.oblata var.alba,respectively.SoLIM expression peaked earlier than the D-limonene emissions in the diurnal experiments,but occurred later when D-limonene had peaked during the flowering phase,indicating that differences in SoLIM gene expression and D-limonene emissions existed.The synthesis of floral scent is thus associated with diverse regulatory mechanisms that require further investigation.
基金supported by the National Natural Science Foundation of China(31870519,31901503 and 31671731)the National Key Research and Development Program of China(2016YFD0101300 and 2016YFD0100202-10)+1 种基金the China Agriculture Research System(CARS-12)the Natural Science Foundation of Jiangsu Province(BK20190267)。
文摘With the increasing promotion of simplified rapeseed cultivation in recent years,the development of cultivars with high resistance to herbicides is urgently needed.We previously developed M342,which shows sulfonylurea herbicide resistance,by targeting acetohydroxyacid synthase(AHAS),a key enzyme in branched-chain amino acid synthesis.In the present study,we used a progeny line derived from M342 for an additional round of ethyl methane sulfonate mutagenesis,yielding the novel mutant DS3,which harbored two mutations in AHAS genes and showed high sulfonylurea resistance.One mutation was the substitution Trp574 Leu,as in M342,according to Arabidopsis protein sequencing.The other site was a newly recognized substitution,Pro197 Leu.A KASP marker targeting Pro197 Leu was developed and reliably predicted the response to sulfonylurea herbicides in the F2 population.The combination of Trp574 Leu and Pro197 Leu in DS3 produced a synergistic effect that greatly increased herbicide resistance.Analysis of the protein structures of AHAS1 and AHAS3 in wild-type and single-gene mutant plants revealed three-dimensional protein conformational changes that could account for differences in herbicide resistance characteristics including toxicity tolerance,AHAS enzyme activity,and AHAS gene expression.
基金supported by the Agricultural Science and Technology Innovation Program, China (ASTIP-TRIC01)
文摘Chalcone synthases （CHS, EC 2.3.1.74） are key enzymes that catalyze the first committed step in flavonoid biosynthesis. In this study, we isolated a chalcone synthase, named NtCHS6, from Nicotiana tabacum. This synthase shared high homology with the NSCHSL （Y14507） gene and contained most of the conserved active sites that are in CHS proteins. The phylogenetic analysis suggested that NtCHS6 protein shared a large genetic distance with other Solanaceae CHS proteins and was the most closely-related to an uncharacterized CHS from Solanum lycopersicum. The expression analysis indicated that NtCHS6 was abundantly expressed in leaves, especially in mature leaves. By scrutinizing its upstream promoter sequences, multiple cis-regulatory elements involved in light and drought responsive were detected. Furthermore, NtCHS6 expression decreased significantly under dark treatment and increased significantly under drought stress suggested that NtCHS6 expression exhibited both light responsiveness and drought responsiveness, and important roles in ultraviolet protection and drought tolerance. Our results might play
基金supported by the National High-Tech R&D Program(863)of China(No.22011AA100605)
文摘Isochorismate synthase(ICS) is a crucial enzyme in the salicylic acid(SA) synthesis pathway. The full-length complementary DNA(cDNA) sequence of the ICS gene was isolated from Artemisia annua L. The gene, named AaICS1, contained a 1710-bp open reading frame, which encoded a protein with 570 amino acids. Bioinformatics and comparative study revealed that the polypeptide protein of AaICS1 had high homology with ICSs from other plant species. Southern blot analysis suggested that AaICS1 might be a single-copy gene. Analysis of the 1470-bp promoter of AaICS1 identified distinct cis-acting regulatory elements, including TC-rich repeats, MYB binding site(MBS), and TCA-elements. An analysis of AaICS1 transcript levels in multifarious tissues of A. annua using quantitative real-time polymerase chain reaction(qRT-PCR) showed that old leaves had the highest transcription levels. AaICS1 was up-regulated under wounding, drought, salinity, and SA treatments. This was corroborated by the presence of the predicted cis-acting elements in the promoter region of AaICS1. Overexpressing transgenic plants and RNA interference transgenic lines of AaICS1 were generated and their expression was compared. High-performance liquid chromatography(HPLC) results from leaf tissue of transgenic A. annua showed an increase in artemisinin content in the overexpressing plants. These results confirm that AaICS1 is involved in the isochorismate pathway.
基金The National Natural Science Foundation of China under contract Nos 40232021 and 40576063
文摘The study aims to reveal phylogenetic and evolutionary relationship between aerobic anoxygenic phototrophic bacteria（AAnPB） and their relatives,anaerobic anoxygenic phototrophic bacteria（AnAnPB） and nonphototrophic bacteria（NPB,which had high homology of 16S rDNA gene with AAnPB and fell into the same genus）,and validate reliability and usefulness of farnesyl pyrophosphate synthase（FPPS） gene for the phylogenetic determination.FPPS genes with our modified primers and 16S rDNA genes with general primers,were amplified and sequenced or retrieved from GenBank database.In contrast to 16S rDNA gene phylogenetic tree,AAnPB were grouped into two clusters and one branch alone with no intermingling with NPB and AnAnPB in the tree constructed on FPPS.One branch of AAnPB,in both trees,was located closer to outgroup species than AnAnPB,which implicated that some AAnPB would be diverged earlier in FPPS evolutionary history than AnAnPB and NPB.Some AAnPB and NPB were closer located in both trees and this suggested that they were the closer relatives than AnAnPB.Combination codon usage in FPPS with phylogenetic analysis,the results indicates that FPPS gene and 16S rRNA gene have similar evolutionary pattern but the former seems to be more reliable and useful in determining the phylogenic and evolutionary relationship between AAnPB and their relatives.This is the first attempt to use a molecular marker beside 16S rRNA gene for studying the phylogeny of AAnPB,and the study may also be helpful in understanding the evolutionary relationship among phototrophic microbes and the trends of photosynthetic genes transfer.
