Auxin response factor(ARF)is a class of transcription factors with B3 domain,which is a direct molecule regulating auxin response and controlling gene expression.A gene,ZmARF10,which encoded ARF protein and actively participated in drought-rehydration stress response,was previously screened in maize by analyzed transcriptome data.To further research the molecular mechanism of ZmARF10 regulating drought resistance of maize,and also provide a new idea for molecular breeding of drought resistance,the gene was firstly analyzed by bioinformatics software.Secondly,Quantitative Real-time polymerase chain reaction(qRT-PCR)was used to detect the expression pattern of ZmARF10 in different tissues,under high temperature,drought,high salt,ABA and restoring treatments,and in different inbred lines.Finally,the function of ZmARF10 was analyzed using CRISPR/Cas9 technology.The results showed that ZmARF10 was located on chromosome 3 of maize,with a total length of 2 127 bp,and encoded 708 amino acids with a typical B3 domain.The upstream 2 kb region of ATG of this gene contained response elements related to methyl jasmonate,auxin,abscisic acid and low temperature.The phylogenetic tree showed that the protein encoded by the ZmARF10 gene was closely related to sorghum.qRT-PCR results showed that ZmARF10 was a constitutive expression gene,and the expression level of ZmARF10 was the highest in mature corn roots of maize.Under high temperature,drought,high salt and ABA treatments,the expression of this gene was significantly up-regulated,and the up-regulation ratio was up to 8.2 times after drought stress.After drought stress,the expression level of ZmARF10 gene was significantly higher in the drought-resistant inbred line Zheng 36 than that in the drought-sensitive inbred line B73.Investigation of Arabidopsis wild type and ARF10-deficient mutants showed that,compared with wild type,the mutant plants showed leaf wilting and even dry death,roots curled,root branch number decreased,and lateral root growth and development were hindered under drought conditions.Determination of physiological and biochemical indexes showed that the relative water content,chlorophyll content and net photosynthetic rate of the deficient mutants were significantly lower than those of the wild-type plants after drought stress,indicating that the drought resistance of Arabidopsis decreased after ARF10 gene was knocked out.
Photoperiod is one of important environmental factors affecting plant bolting and flowering which are regulated by plant phytochrome proteins.The signal network mediated by PHYB has an important inhibitory effect on plant bolting and flowering.Previous studies revealed that there were large segment insertion/deletion differences between the PHYB gene promoter of the Chinese cabbage late-bolting line 06-247 and the easy-bolting line He102.In order to further investigate the impact of promoter mutation on PHYB and the key genes of its downstream pathways,this study was conducted.Based on bioinformatics methods,the redundancy characteristics of phytochrome genes in genome Chinese cabbage were firstly analyzed.It was found that the Chinese cabbage genome contained six phytochrome genes,of which PHYA had two copies,and PHYB,PHYC,PHYD,and PHYE all had only one copy.Then amino acid sequence alignment was used for screening of the specific sequence of PHYB.Antigenic determination clusters were designed based on the specific sequence and the antibodies against PHYB was prepared.The fluorescence quantitative RT-PCR technology and Western Blot technology were used to study the relative content of PHYB in 06-247 and He102.At the same time,the dynamic changes of key regulatory genes such as CCA1,FLC,CO and FT which in the downstream of PHYB pathway were also compared.The results showed that the promoter mutation caused significant differences both in level of mRNA in 06-247 and then significantly increased the protein level of PHYB.At the same time,the downstream regulatory genes such as CCA1,FLC,CO and FT were highly expressed in 06-247,which had an important impact on bolting resistance of 06-247.
To reveal the distribution preferences of SSR in foxtail millet genome,the study analyzed distribution regularities of SSRs among the 5'-flanking regions,the 3'-flanking regions,the extron regions and the intron regions in gene locus from foxtail millet genome using bioinformatics methods.The results showed that the relative frequency,a ratio of the frequency of a type of SSR in a certain genomic region by the frequency of this SSR in whole genome,of SSRs decreased with increasing of the distance of 5'-flanking regions from start codon,and was more than 1.50 obviously in a range from 1 bp to 700 bp.However,the relative frequency of SSRs was close to 1.00 in each of other three regions.Furthermore,with increasing of the distance of 5'-flanking regions from start codon,the relative frequency of various SSR types revealed the various changing trends,and as a whole,6 SSR types,including CCG,AG,AGG,AGGG,ACC and AGC,showed both the high frequency and the high relative frequency,and of these 6 types,the distance range(1—1 800 bp)of AG,in which its relative frequency was more than 1.50,is the maximum while that of CCG is minimal(1—200 bp).Meanwhile,in the extron regions,5 SSR types with high C/G contents,including CCG,AGG,AGC,ACG and ACC,showed both a high frequency and a high relative frequency.It was concluded that SSRs distributed preferentially in a length range from about 1—700 bp of 5'-flanking regions with 6 main specific SSR types,including CCG,AG,AGG,AGGG,ACC and AGC;on the other hand,in 3'-flanking regions,the intron regions and the extron region,distribution of SSR is close to that in genome,and however,in the extron region,a total of 5 SSR types,including CCG,AGG,AGC,ACG and ACC,are obviously specifically distributed.
Leaf color mutant is an ideal material for studying C4 photosynthesis pathway and chlorophyll metabolism mechanism.In order to study the molecular mechanism of yellow-green leaf mutation in millet,and lay a foundation for the functional study of yellow-green leaf genes and the molecular mechanism analysis of chlorophyll metabolism,a stable hereditary yellow-green leaf mutant ygl7 was identified in the ethyl methacrylate(EMS)mutant library of Changnong 35.Agronomic traits,photosynthetic pigment content and photosynthetic parameters,and chloroplast ultrastructure observation were carried out on the mutant and wild type.At the same time,genetic analysis of mutant leaf color was performed,primary mapping was performed by BSA method,fine mapping was performed by an F2 population,and candidate genes were predicted according to functional annotation combined with RNA-Seq.The expression pattern was analyzed by qRT-PCR,and the protein interaction was verified by yeast two-hybrid experiment.The results showed that the leaves of ygl7 were obviously yellow-green at seedling stage and elongation stage,and gradually turned to light green at heading stage compared with the wild type.The chlorophyll content and photosynthetic rate of ygl7 during the whole growth stage were significantly lower than that of the wild type,and the chloroplast structure was abnormal.Genetic analysis showed that ygl7 yellow-green leaf phenotype was controlled by a pair of recessive single genes.Yellow-green leaf gene was located in the 434.9 kb region of chromosome Ⅶ.Candidate gene analysis predicted that Seita.7G290300 encoding protoporphyrin Ⅸ magnesium chelatase Ⅰ was the candidate gene for regulating yellow-green leaf.The results of qRT-PCR showed that Seita.7G290300 was highly expressed in leaves,and the expression of Seita.7G290300 in mutant was lower than that of wild type.The expression levels of genes related to chlorophyll synthesis pathway(CHLD,CHLI)and photosystem(LHCB1,LHCB6)were down-regulated in the mutants.The experiment of yeast two-hybrid showed that SiYGL7 interacted with MORF2.
To improve seedling blast resistance of rice cytoplasmic male sterile(CMS)line Taonong1A and its maintainer Taonong1B,the Chinese local rice variety Gumei 4 was used as the donor parent harboring the broad-spectrum and durable blast resistance gene Pigm,and the Pigm-tightly-linked codominant marker T9E3 was developed and employed in marker-assisted selection(MAS)breeding practice.T9E3 showed clear and stable polymorphism between the donor and acceptor parents,amplifying a 926 bp DNA band from Gumei 4 genome,while the PCR product from either genome of Taonong1B or Taonong1A was about 2 000 bp.Greenhouse inoculation assay was performed using thirty-two Magnaporthe oryzae isolates collected from different rice areas,Gumei 4 and the improved rice lines showed much higher resistance frequency(90.63%)than that of two acceptor parents Taonong1B and Taonong1A(9.38%).Accordingly,Gumei 4 and the improved rice lines showed high-level seedling blast resistance(0 grade)but highly susceptible(8 grade)for two acceptor parents in natural nursery.Three blast resistant CMS pure lines(Taonong1A-Pigm-1—Taonong1A-Pigm-3)and the corresponding resistant maintainers(Taonong1B-Pigm-1—Taonong1B-Pigm-3)were preliminarily bred through MAS breeding,and one pair of them,Taonong1A-Pigm-2/Taonong1B-Pigm-2,was screened for potential application duo to the high-level blast resistance and elite sterility,agronomic and yield traits.Taonong1A-Pigm-2 showed complete male sterility with nearly 60% typical abortive pollens and 40% round abortive pollens,higher stigma exsertion rate(71.1%)and lower enclosed spikelet rate(33.6%),Taonong1B-Pigm-2 exhibited excellent agronomic and yield trait indexes,such as moderate sowing-heading duration(69 days),plant height(76.2 cm),main panicle length(28.1 cm),panicles per plant(15.8),spikelets per panicle(115.3)and 1000-grain weight(27.7 g)。New parent materials were created for the application of cytoplasmic male sterile(CMS)line hybrid rice breeding,and the comprehensive improvement of seedling blast resistance and characters of Taonong1A and Taonong1B were realized.
Obtaining the LaeA gene sequence of Ganoderma lucidum through electronic cloning,analyzing its gene sequence information and preliminarily exploring its regulatory function.This study adopts the method of electronic cloning,using the known LaeA protein sequence of Penicillium citrinum as a template,perform sequence similarity search and alignment(Blast)in the EST database of Ganoderma lucidum and obtain the cDNA sequence of the LaeA gene of Ganoderma lucidum through electronic cloning methods such as sequence splicing,sequence validation and sequence extension.Some characters of amino acids encoded by LaeA gene,including the physical and chemical properties,hydrophobicity/hydrophilicity,subcellular localization,secondary and tertiary structure of protein,and phylogenetic relationship were analyzed by bioinformatics tools.The length of LaeA gene from G.lucidum was 1 134 bp,encoded 378 amino acids.The protein encoded molecular weight of 42.895 3 ku.The protein was an instability protein that was present in the cytoplasm and not secreted to the extracellular.Furthermore,the structure of LaeA protein was mainly composed by 47.88% random coil,33.33%α-helix and 18.78% extension strand,contained SAM binding site,belonged to AdoMet_MTases superfamily proteins.Phylogenetic analysis showed that LaeA was closely related to white rot basidiomycetes such as Trametes versicolor and Dichomitus squalens;the Real-time Fluorescence Quantitative PCR results showed that the expression level of LaeA in Ganoderma lucidum cells during liquid static culture was significantly higher than in oscillatory culture.It is speculated that the LaeA protein as a methyltransferase protein that participates in histone methylation modification,thereby affecting the expression level of gene clusters.
To investigate the nature and function of PlSPL1 (SPL)gene of Paeonia lactiflora,further elucidate the commonalities and characteristic differences of the PlSPL1 gene in different species,and explore the role of PlSPL1 in the degree of stem straightening in Paeonia lactiflora. The full-length sequence of the PlSPL1 gene was obtained by RACE technology using the stalk of herbaceous peony Hongfeng as the research material,and the structural,physicochemical properties,and phylogenetic relationships of PlSPL1 were analyzed and predicted using bioinformatics software,then,the expression level of PlSPL1 in different developmental stages of herbaceous peony stems was analyzed using qRT-PCR technology,and subcellular localization analysis of the protein was conducted using laser confocal microscopy technology.The results showed that the open reading frame of the PlSPL1 gene was 3 000 bp,encoding 999 amino acids.The protein had a molecular formula of C4869H7682N1406O1497S43,a molecular weight of 111.25 ku,a theoretical isoelectric point of 6.26,encoding a hydrophilic unstable acidic protein,phosphorylation modification predominantly on serine,no signal peptide,with transmembrane structure,and had a secondary structure consisting mainly of random coil.Phylogenetic tree analysis revealed that the PlSPL1 protein was most closely related to peony,followed by a closer relationship with grape;protein sequence comparison analysis revealed that the PlSPL1 protein had a conserved SBP domain which was unique to SPL transcription factor family.The relative expression analysis found that PlSPL1 gradually showed a decreasing trend with stem development,indicating that PlSPL1 was negatively regulating the stem development of herbaceous peony and it was hypothesised that PlSPL1 played important roles in degree of stem straightness;and the subcellular localisation showed that the PlSPL1 protein was localised in the nucleus.The above results indicate that PlSPL1 is participated in the stalk development of herbaceous peony.
In order to clarify the sequence characteristics of the MYB62 transcription factor,expression changes after stress,and further explore the biological function of the MYB62 transcription factor,kohlrabis were used as plant materials.MYB62 transcription factors were obtained by homologous cloning method and bioinformatics was analyzed.Spatiotemporal expression analysis of MYB62 and its expression after stress were performed by Real-time PCR.Results of gene cloning showed that gDNA length of BocMYB62 gene was 1 353 bp,the length of CDS was 837 bp,which contained four exons and three introns and encoded 278 amino acids.The sequence structure analysis showed that BocMYB62 was a hydrophilic protein with two SANT-MYB domains,belonging to the R2R3-MYB type in the MYB transcription factor family.The spatial structure prediction revealed a typical α-helix structure.Phylogenetic analysis indicated that BocMYB62 was closely related to MYB62 in Brassica napus.The spatiotemporal expression results showed that BocMYB62 was consistently higher in green kohlrabi than that in purple with clear tissue specificity.The expression of BocMYB62 was significantly increased during drought stress with the highest expression under 12 h.The BocMYB62 expression was significantly lower than control and the lowest at 4 ℃ cold stress.These results inferred that BocMYB62 may be involved in the regulation of anthocyanin biosynthesis and the regulation of stress.It provides a theoretical basis for the subsequent functional identification of MYB62 transcription factors.
The bioinformatic analysis of SAUR genes aims to unveil their structure,function,and evolutionary relationships.Simultaneously,through subcellular localization,the goal was to understand their position within the cell,thereby inferring their biological functions.Such analyses contribute to a deeper understanding of the gene's mechanistic role and potential applications within an organism.This held particular theoretical and practical significance for regulating the germination of windproof seeds,reducing seed dormancy duration,and facilitating green cultivation and breeding of wind-resistant plants.In order to explore the biological characteristics and functions of SAUR genes,SdSAUR32 and SdSAUR23 were cloned from Saposhnikovia divaricata,and bioinformatics analysis was conducted to analyze the expression and subcellular localization of these two genes in different dormancy stages of Saposhnikovia divaricata seeds.The findings indicated that the SdSAUR32 protein had a molecular weight of approximately 14.48 ku(pI 6.45),while the SdSAUR23 protein had a molecular weight of around 10.29 ku(pI 8.00),confirming that both proteins were hydrophilic.The conserved domain analysis of SdSAURs gene encoded protein showed that SdSAUR32 and SdSAUR23 had the same conserved domain,which belonged to auxin induced superfamily members.The phylogenetic analysis showed that both SdSAUR32 and SdSAUR23 encoded protein was the closest relative to Daucus carota. The subcellular localization results indicated that both SdSAUR32 and SdSAUR23 were found within the nucleus.The SdSAUR32 and SdSAUR23 genes were found to be expressed during different stages of Saposhnikovia divaricata seed development.It was observed that the expression levels of these two genes were significantly higher during the germinal stage compared to the release dormancy stage and dormancy stage.This suggested that SdSAUR32 and SdSAUR23 may have a crucial role in the germination,growth,and development of Saposhnikovia divaricata seeds.This study offers not only insights for investigating the role of the SAUR gene family but also establishes atheoretical framework for examining the function of other genes in the parsnip family at the molecular level.
In order to study the correlation between stem traits and the formation of elasticity and density tolerance of maize,and further reveal the mechanism of lodging resistance,6 maize varieties with different lodging resistance were selected as materials,and 3 planting densities of 6.0×104,7.5×104 and 9.0×104 plants/ha were set.The stem pull lodging angle in the field was used as the evaluation index of elasticity,and the morphological characteristics between plants and basal nodes.The internode anatomical structure,material accumulation and mechanical characteristics were analyzed.The results showed that plant height,ear height,basal internode length,thickness,epidermal thickness,crusty tissue thickness,total vascular bundle,sheath area of small vascular bundle,fresh weight per unit length,dry weight per unit length and content of each component,puncture and folding strength had significant effects on the stem pull lodging angle,among which internode diameter(r=0.521**)and dry weight per unit length(r=0.562**)had the greatest effects.The greater the planting density,the smaller the internode diameter,the thickness of the hard skin tissue,the total number of vascular bundles,the fresh weight per unit length,the dry weight,the content of each component,the lodging resistance,the worse the elasticity of the stem.There were significant differences in stem traits among different varieties.The elasticity related traits of Lishou 1,Chuangyu 107,Jingnongke 728 and MC278 were better than those of other varieties,and the variation amplitude was smaller with the increase of density,so the elasticity and density tolerance were stronger.Traits such as internode diameter and dry weight per unit length had significant effects on stem pull lodging angle, that was stalk elasticity,and the variation amplitude of these traits after densification determined the density tolerance of stalk.
To clarify the effects of drought stress and cultivars on the net photosynthetic rate(Pn),the activities of photosynthetic enzymes in flag leaf in the afternoon(FLA)during grain filling stage and grain yield of winter wheat,a pond culture experiment with four water levels and two winter wheat cultivars was conducted under the condition of rainproof pond cultivation in 2019—2021.The four water treatments included severe drought(W1),moderate drought(W2),mild drought(W3),and suitable water supply(W4).The Pn,and the activities of Rubisco,RCA,PEPC,ATPase and PPDK in flag leaf from 14:00 to 16:00 during the early(EGFS)and medium(MGFS)grain filling stage,and the grain yield at maturity of strong drought resistance cultivar Jinmai 47(JM47)and weak drought resistance cultivar Yanzhan 4110(YZ4110)were investigated.Drought stress decreased the FLA Pn and activities of most photosynthetic enzymes,and the grain yield of wheat.The decrease of these indexes increased with drought stress degree,but the effects were different among varieties and years.Compared with W4,the FLA Pn of JM47 under W1,W2 and W3 decreased by 33.6%—40.6%,12.0%—30.5% and 5.0%—13.5%,as well as YZ4110 decreased by 44.0%—52.0%,22.5%—38.1% and 11.5%—20.5%,respectively.Compared with W4,the FLA Rubisco activity decreased during EGFS but increased during MGFS for JM47,while it decreased by 13.3%—25.6%,7.1%—14.0% and 11.2%—11.6% for YZ4110,respectively,under W1,W2 and W3.Compared with W4,the FLA RCA activity significantly decreased under most drought treatments during EGFS,while increased under W2 and W3 for JM47 and decreased under W1 and W2 for YZ4110 during MGFS.Compared with W4,the FLA ATPase activity of JM47decreased under W1 but increased under W3,while that of YZ4110 decreased under W1,W2 and W3 by 19.3%—48.7%,7.2%—24.2% and 0.1%—8.9%,respectively.The FLA PEPC activity under different treatments varied with growing seasons and varieties.Compared with W4,the FLA PPDK activities of JM47 and YZ4110 under W1 were decreased by 12.4%—18.8% and 16.7%—18.2%.Compared with YZ4110,in most conditions,the FLA Pn and photosynthetic enzyme activities of JM47 had no significant difference under suitable water supply(W4),but increased under drought treatments(W1,W2 and W3).The results of correlation analysis showed that yield,FLA Pn were significantly positively correlated to FLA ATPase activity during EGFS and MGFS,as well as FLA PEPC activity during EGFS.Thus,increasing the FLA ATPase and PEPC activities during the grain filling period is conducive to the increase of FLA Pn and grain yield of wheat.
Fruit development is the key stage which determines the yield and quality of grape.WRKY family transcription factors play important roles in regulating plant development and environment adaptation.Ethylene is the important plant hormone which participates in regulation of fruit development,and ACC synthase is the key enzyme that limits ethylene synthesis.Using grape variety Zuoyouhong, VvWRKY13 overexpressing grape callus,as well as VvWRKY13 heterologous over-expressed tomato lines as materials,the role of VvWRKY13 from grape in fruit development and its relationship with ethylene were studied though plant physiological and biochemical methods as well as molecular biological techniques.The results displayed that the expressions of VvWRKY13 as well as ACC synthase genes VvACS2 and VvACS7 significantly upregulated at early stage of fruit development,and in VvWRKY13 over-expression grape callus,the expression of VvACS7C was significantly higher than control,but the expression of VvACS2 displayed no significant difference compared with control.Yeast one hybrid experiment showed that VvWRKY13 could directly bind to the VvACS7 promoter,VvWRKY13 had no direct interaction with VvACS2.We also found that the ethylene content and the expression of ACS family members,such as SlACS1b,SlACS4 and SlACS6,the ethylene synthesis genes,were significantly induced in VvWRKY13 heterologous over-expressed tomato,and the time from flowering to breaking of tomato was 3—6 days shorter than wild type;the above results indicated that,VvWRKY13 could regulate ethylene synthesis by promoting the expression of ACC synthase gene to participate in regulation of fruit development.
Knockout mutants ossaur55-1 and ossaur55-2 obtained by CRISPR/Cas9 technology in Francis background were used as research objects to study whether they affect the changes of agronomic traits such as plant type,leaf color,ear weight per plant and seed setting rate of rice.To explore whether OsSAUR55 regulates rice plant type by participating in GA pathway,and to provide theoretical basis for regulating rice plant type by related hormone pathway.The main agronomic traits of wild type(WT)and knockout mutant in adult plant stage were examined,including plant height,ear length,ear weight per plant,ten grain length,ten grain width,blade length,and blade width.Chlorophyll content and net photosynthetic rate of wild type and mutant leaf were measured by chlorophyll meter and photosynthetic rate meter.Real-time Fluorescence Quantitative PCR(qPCR)was used to determine the expression of OsSAUR55 gene in different tissues and different time points of rice seed germination.The subcellular localization of OsSAUR55 protein was observed by prokaryotic expression method.The contents of endogenous hormones(GA and IAA)in wild type and mutant were determined by liquid chromatography-tandem mass spectrometry(LC-MS/MS).The expression of genes related to GA pathway was further analyzed.In the adult plant stage,mutant ossaur55-1 and ossaur55-2 showed significant phenotypic changes,in which the mutant was significantly shorter in plant height,greener in leaf color,increased in chlorophyll content,decreased in photosynthetic efficiency,decreased in seed setting rate,decreased in 1000-grain weight and decreased in panicle weight per plant than the wild type.The quantitative results showed that OsSAUR55 expression was the highest in the plumule,but not in the flag leaf.Confocal results showed that OsSAUR55 may be localized in cell membrane and nucleus.The results of hormone detection showed that the content of GA4 was decreased significantly.Gene expression analysis showed that the expression levels of genes KAO,GA13ox1,GA20ox1 and KO1 in the GA synthesis pathway were significantly higher than those of the wild type,and the expression levels of GA2ox5,GA2ox8 and GA2ox9 in the GA metabolic pathway were also significantly higher than those of the wild type.It is preliminarily speculated that the deletion of OsSAUR55 gene led to the decrease of endogenous GA4 content,which may regulate rice plant type through GA pathway,and then affect rice yield.
To investigate the effect of exogenous melatonin (MT) on salt tolerance of soybean seedlings,and to screen the appropriate application concentration under different salt stress.The soybean variety Tianyou-2986 was used as the test material,and 3 salt concentrations (low salt S3:3 g/L,medium salt S5:5 g/L,high salt S7:7 g/L ) and 6 MT concentrations were set(M0:0 μmol/L,M1:25 μmol/L,M2:50 μmol/L,M3:75 μmol/L,M4:100 μmol/L,M5:150 μmol/L),the morphological parameters,biomass,antioxidant enzyme activity and osmoregulatory substance content of soybean seedlings were analyzed.With the increase of salt stress,the root morphological parameters,biomass,root-shoot ratio,antioxidant enzyme activity and osmoregulatory substance content of soybean seedlings decreased,while the malondialdehyde content increased.Under low salt (S3) and medium salt (S5) stress,The number of total length,lateral roots,SOD and POD of 50 μmol/L MT were increased by 52.30%,19.98%,74.10%,40.03% (low salt) and 68.52%,19.24%,81.72% and 37.42% (medium salt),respectively.Under high stress (S7),75 μmol/L MT increased by 71.17%,19.11%,80.79% and 27.01%,respectively.Under salt stress,exogenous 25—100 μmol/L MT promoted soybean seedling growth and improved salt tolerance to different degrees.The overall evaluation of the affiliation function showed that 50 μmol/L MT was the most effective in alleviating salt damage under low and medium salt stress,and the suitable concentration of MT was 75 μmol/L under high salt stress,and the main reason for the alleviation of salt damage in soybean seedlings was that MT increased the activities of antioxidant enzymes and osmoregulatory substance content,and reduced the content of malondialdehyde,which could alleviate oxidative and osmotic stresses of soybeans under salt stress.
To clear under the condition of slow release nitrogen fertilizer applying different content of maize development rule of the diurnal change of photosynthetic characteristics and growth,the light response curve was to fit,so as to farming-pastoral zone of spring maize cultivation and nitrogen utilization to provide theoretical basis.This study took Guangde 5 as the research object,and based on the long-term positioning experiment in 2018,the leaves ear position in spinning—tasseling stage of maize were mesured and analyzed under six nitrogen gradient of N 0(N0,CK), 120(N8), 180(N12), 240(N16), 300(N20),360(N24) kg/ha in 2019 and 2020. SPAD value, intercellular carbon dioxide concentration, stomatal conductivity, diurnal variation of net photosynthetic rate, transpiration rate, light response curve and dry matter accumulation rule.The results showed that with the increase of nitrogen fertilizer application, the changes of SPAD value, net photosynthetic rate, transpiration rate and stomatal conduction degree day of maize increased first and then decreased,and N16 treatment was the highest,and intercellular carbon dioxide concentration was the lowest.The changes of net photosynthetic rate,transpiration rate and stomatal conduction degree day showed unimodal curve.The light response curve fitting analysis showed that the maximum net photosynthetic rate of N16 treatment was the highest.In 2019 and 2020,N16 treatments were 37.48%,29.51%,31.85%,18.17%,37.32% and 80.04%,59.73%,50.30%,6.42%,62.51% higher than N0,N8,N12,N20 and N24 treatments,respectively.Dry matter mass accumulation in whole plant and ear were positively correlated with maximum net photosynthetic rate,apparent quantum efficiency and SPAD value.Comprehensive analysis showed that the most suitable nitrogen fertilizer application rate in western Inner Mongolia was 240 kg/ha.
In order to study the effect of applying N,P and K fertilizer on the yield and nutrient use efficiency of oil sunflower under the double cropping pattern of winter wheat-summer oil sunflower in Southern Shanxi,a field plot fertilizer experiment was carried out with Linkui 4 from 2019 to 2021.Four fertilization treatments including NPK,PK,NK and NP were set up.The effects of different fertilization treatments on yield response,agronomic efficiency and fertilizer use efficiency of N,P,K were analyzed.The results showed that the three-year average yield of oil sunflower treated with NPK was 3 634 kg/ha(3 502—3 818 kg/ha),which was 19.6%,12.1%,13.7% higher than that of PK/NK/NP,respectively.PK treatment had the lowest yield.The average yield responses of N,P and K fertilizers were 588.8(481.0—704.7 kg/ha),391.8(296.0—488.7 kg/ha),438.2 kg/ha(329.3—629.7 kg/ha),respectively.The agronomic efficiencies of N,P and K were 3.8(2.8—4.9 kg/kg),3.1(2.1—4.1 kg/kg),2.9 kg/kg(2.2—4.2 kg/kg),respectively.The aboveground accumulations of N,P and K were 174.4(172.5—176.8 kg/ha),60.0(58.3—61.8 kg/ha),241.1 kg/ha(236.2—246.8 kg/ha),respectively.The nutrient use efficiency of N,P,K was 36.3%(36.0%—36.7%),15.7%(15.3%—16.2%)and 47.1%(46.0%—47.9%),respectively,of which K>N>P.So the application of N,P and K fertilizer could not only significantly increase the oil sunflower yield but also the aboveground accumulation of N,P and K.Among the three kinds of NPK fertilizers,N increased the yield most,and K uptake by the aboveground part and utilization rate of oil sunflower was the highest.Therefore,in the production we should pay attention to the reasonable application of N,P and K,while N and K should be supplemented by seed fertilizer and topdressing.
In order to study the differences in yield and nitrogen utilization,transport and accumulation of different winter wheat cultivar(line)types in the Northern Huanghuai,the yield traits and nitrogen utilization efficiency of 29 and 26 winter wheat cultivars(lines)for testing in the Northern Huanghuai wheat area were investigated and analyzed in 2021—2022 and 2022—2023,respectively.Through cluster analysis,wheat was classified into four types:high-yielding,medium-high-yielding,medium-yielding and low-yielding.Nitrogen content of wheat stems,leaves,spikes,and grains at maturity were measured at anthesis and maturity,respectively,to analyze the relationship between parameters related to nitrogen utilization traits and yield.The results showed that the average yields among high-yielding,medium-high-yielding,medium-yielding and low-yielding varieties(lines)differed significantly from 2021 to 2023,with the number of spikes per hectare of high-yielding varieties significantly higher than that of other types.At the anthesis stage,the nitrogen accumulation and distribution rate of each organ were stems and sheaths>leaves>spikes;at the maturity stage,the nitrogen accumulation of each organ was grains>stems and sheaths>spikes>leaves.The post-anthesis nitrogen accumulation,pre-anthesis nitrogen transport and its contribution to the grain of different varieties of wheat were higher in high-yielding varieties(lines),and the pre-anthesis nitrogen transport and its contribution to the grain were greater than the post-anthesis nitrogen accumulation and its contribution to the grain.Nitrogen utilization efficiency,nitrogen harvest index,nitrogen accumulation at anthesis and nitrogen accumulation at maturity were significantly and positively correlated with wheat grain yield.Therefore,water and fertilizer management can be carried out through the nitrogen uptake and transfer pattern and distribution characteristics of different varieties,or selecting and breeding wheat varieties with high nitrogen utilization efficiency to achieve high-yield and high-efficiency production of wheat.
In order to explore the role of RPM1 in sorghum disease resistance,a sorghum SbRPM1 gene was obtained from sorghum smut resistant variety SX44B by homologous cloning method.The bioinformatics analysis results showed that the total length of the cDNA of SbRPM1 gene was 2 802 bp,encoding 933 amino acids,and its protein had a theoretical molecular weight of 106.1 ku and an isoelectric point of 7.11,which was a hydrophilic protein.The SbRPM1 protein had no transmembrane structure,and its subcellular localization was in the cytoplasm.Conservative domain analysis showed that SbRPM1 protein contained RX-CC-like,NB-ARC and LRR domains,and belonged to CNL proteins in the NLRs family.Phylogenetic analysis showed that SbRPM1 protein was most closely related to the RPM1 protein of Miscanthus lutarioriparius.The expression pattern of SbRPM1 gene was detected by Real-time quantitative PCR,and the results showed that the expression of SbRPM1 gene was higher in leaves and inflorescence,followed by roots,and the lowest in stem.The expression of SbRPM1 gene was significantly up-regulated at 24—72 h in disease-resistant varieties after inoculation with Sporisorium reilianum pathogen,suggesting that this gene could be induced by S.reilianum and played an important role in sorghum disease resistance.In this study, the CDS sequence of the SbRPM1 gene was cloned for the first time in sorghum, and the structure, nature and expression of the gene were characterized.
In order to investigate the effect of iron ions on the biological characteristics and pathogenicity of Verticillium dahliae in sunflower,the growth rate,conidial production,number of microsclerotia,crude toxin secretion,cell wall degrading enzymes activity and pathogenicity of V.dahliae were measured after the exogenous addition of iron ions to the culture medium.The results showed that the growth rate and conidial production of mycelium showed an increasing trend after adding different concentrations of iron ions in an exogenous,compared with the control,the growth rate of V.dahliae was the fastest after adding 80 μmol/L of iron ions,and the colony diameter was 68.81 mm with a growth rate of 21.40%;the conidial production was 2.58×107 conidia /mL with a growth rate of 21.13%;the number of microsclerotia also increased with the increase of exogenously added iron ion concentration,and after 5 days of culture,its increase was 51.53% compared with the control.The crude toxin secretion was increased after adding 80 μmol/L of iron ions,and the increase rate was nearly doubled.The activity of cell wall lyase increased with increasing concentration of exogenous iron ions and increased at 80 μmol/L reached the strongest.In addition,the pathogenicity of V.dahliae increased accordingly with the increase in the concentration of iron ions in the medium,as shown by the increase in the disease index from 35.00 to 62.20 when the concentration of iron ions added exogenously from 0 to 80 μmol/L,with a growth rate of 77.71%.In conclusion,the exogenous addition of iron ions not only accelerates the growth of V.dahliae,and promotes conidial production and microsclerotia formation,but also enhances the pathogenicity of V.dahliae.
Luchuan pig myoblast determining gene 1(MyoD1)was cloned,bioinformatics analysis was performed,and its expression was analyzed in different tissues of Luchuan pigs,to explore the role of MyoD1 gene in the growth and development of skeletal muscle.Published MyoD1 gene of wild boar sequence on NCBI as the template,designed specific primer,cloned the CDS region of MyoD1,and compared the similarity with wild boar,cattle,sheep,human,mouse and rat gene sequence,constructed system tree for bioinformatics analysis by online software.Then,the relative expression of MyoD1 gene in different tissues of Luchuan pigs by RT-PCR.The CDS of MyoD1 gene in Luchuan pig was 960 bp,encoding 319 amino acids,five base mutations.The similarities of MyoD1 genes between Luchuan pig and wild boar,cattle,sheep,human,mouse and rats were 99.2%,93.0%,92.3%,90.0%,84.3% and 84.0%,respectively.Total number of MyoD1 gene atoms in Luchuan pigs was 4 680,molecular mass of the protein was 33.99 ku,molecular formula was C1457H2296N442O471S14;the instability coefficient was 63.87,indicating a lack of stability;isoelectric point was 5.63,which belongs to the acidic protein;no transmembrane structure existed,found 35 phosphorylation sites and 1 glycosylation site;protein secondary structure was dominated by an irregular coil,which accounting for 60.69%.The MyoD1 gene in Luchuan pigs had the highest expression in the longest dorsal muscle,which was significantly higher than in other tissues,while kidney had the lowest expression.MyoD1 gene was expressed in all tissues of Luchuan pigs and mainly in the longest dorsal muscle,we speculated that MyoD1 gene might play a crucial role in the muscle growth and development of Luchuan pigs.
This study aims to obtain the MFSD4A gene sequence in Maiwa yak and elucidate its expression patterns in different tissues,analyze the mRNA expression levels of the interacting proteins of MFSD4A,and determine the subcellular localization of the MFSD4A protein,and provide a theoretical basis for further research on regulating MFSD4A in the growth and development of yak testis.RT-PCR was used to obtain the MFSD4A gene CDS sequence in Maiwa yak and perform bioinformatics analysis.RT-qPCR was used to detect the expression differences of the MFSD4A gene in 10 tissues,including heart,liver,spleen,lung,kidney,gluteal muscle,back muscle,fat,testis,and lymph,and the correlation between the interacting proteins and MFSD4A.pEGFP-MFSD4A fusion plasmid was transfected into Madin-Darby bovine kidney cells(MDBK)to investigate the subcellular localization of the MFSD4A protein.The results indicated that the entire length of the MFSD4A gene CDS region in the macaque yak was 1 530 bp,encoding 509 amino acids with a theoretical isoelectric point(PI)of 8.66.It had 12 transmembrane domains and belonged to the alkaline transmembrane protein.RT-qPCR results showed differential expression of MFSD4A mRNA in various tissues of the macaque yak,with the highest expression in the testis.It was also highly positively correlated with the expression levels of MFSD9,CBY1,INHBA,UNC93A,SVOP,and ID1 in the testis,suggesting that the MFSD4A gene might be related to the regulation of growth and development of yak testis.Fluorescence localization of the pEGFP-MFSD4A fusion vector transfected into Madin-Darby bovine kidney cells(MDBK)showed that the protein was mainly located in the nucleus.
In order to explore the expression and function of interferon-stimulated exonuclease gene 20(ISG20)in the formation of receptivity in beef cattle endometrial epithelial cells.The CDS region of the beef cattle ISG20 was obtained by RT-PCR combined with sequencing.The sequence analysis of the ISG20 and its encoded proteins was performed using bioinformatics online tools.The expression of the ISG20 and its related genes was detected by qRT-PCR in the formation of receptivity in beef cattle endometrial epithelial cells.The results showed that the length of CDS region of the ISG20 was 516 bp,which could encode a single protein consisting of 176 amino acids with no transmembrane domain and unstable basic hydrophilicity.The protein contained the DEDD(or DnaQ-like)exonuclease superfamily domain,interacted with 10 proteins,including ISG15 and MX2,and functioned mainly in the cytoplasm.In addition,the qRT-PCR results showed that compared with the control group,the expression of the ISG20 and its related ISG15 and MX2 was both significantly up-regulated in the formation of receptivity in beef cattle endometrial epithelial cells induced by progesterone(P4)-co-interferon-tau(IFN-τ).ISG20 may be involved in the formation of receptivity in beef cattle endometrial epithelial cells and play a role in biological processes such as early pregnancy.
In order to study the structure and function of DJ-1 gene in yaks,this experiment used Meiren yak adipose tissue cDNA as a template,cloned the coding sequence (CDS) of yak DJ-1 gene using RT-PCR,and performed bioinformatics analysis,including domain prediction and protein physicochemical properties prediction of the gene sequence.Then,tissue expression analysis was performed using RT-qPCR.The results showed that the CDS region of the DJ-1 gene in Meiren yak had a total length of 570 bp and encoded 189 amino acids;the prediction of the DJ-1 domain in yaks showed that there was one PfpI domain containing the Pfam in positions 29—168 of the DJ-1 amino acid sequence.By predicting structure and function of the DJ-1 protein,the results showed no transmembrane structure and no signal peptide region;molecular weight was 20.035 31 ku,total number of atoms was 2 870,theoretical isoelectric point was 6.84,instability coefficient was 28.37,and expressed as a stable protein;the fat coefficient was 101.11,with a total average hydrophilicity coefficient of -0.004.It was a hydrophilic protein with a total of 11 phosphorylation sites.Meanwhile,the results of subcellular localization prediction indicated that the DJ-1 protein of Meiren yak was mainly distributed in the cytoplasm and mitochondria.The phylogenetic tree showed that Meiren yak had the closest genetic relationship with wild yaks and European cattle,and the farthest genetic relationship with Arctic foxes and wide snouted manatees;the RT-qPCR results showed that it was expressed in the heart,liver,spleen,lungs,muscle,fat,and testicular tissues of adult Meiren yaks,with the highest expression level in the heart and muscle tissues,and the lowest expression level in the liver and testicular tissues.
The aim of this study was to analyze the regulatory effect of RXFP2 gene on horned phenotype and related SNP markers in horned and hornless Oula sheep populations,in order to provide technical support for genetic improvement and breed breeding of Oula sheep.DNA was extracted from 100 ewe blood samples of healthy adult Oula sheep in Henan County,Qinghai Province,including 50 blood samples with horns and 50 blood samples without horns,respectively,15 samples with and without horns were randomly selected for whole genome sequencing,and then all blood samples were amplified by multiple PCR to verify the SNPs of RXFP2 gene.Genome-wide detection results showed that there were strong selection signals in chromosome 10 of Oula sheep,including RXFP2 gene,and the strongest selection signals appeared in the RXFP2 gene segment.Multiple PCR tests verified the accuracy of the 4 coding SNPs screened by whole genome resequencing.Seven high frequency SNPs in the intron region of RXFP2 gene were detected in Oula sheep(29508704G>A,29509428A>C,29509766G>A,29512170G>A,2951276G>A,29514968T>A,29521377C>A).The genotypes of the 7 SNP loci were separated in hornless phenotypes.100% of homozygous genotypes appeared horned or hornless trait,89.66% of heterozygous genotypes were hornless trait and 10.34% were horned trait.The frequency of mutant alleles was less than that of wild alleles,and the population showed an unbalanced state of Hardy-Weinberg,which meant subject to high artificial selection intensity,but the content of polymorphic information was moderate,and the potential of breeding and improvement was still large.In conclusion,the present study confirmed the strong regulatory effect of RXFP2 gene on horn trait in Oula sheep,and filtered out 7 molecular markers of RXFP2 gene on horn trait in Oula sheep population.The relevant results can be used as molecular markers for genetic improvement of the hornless population of Oula sheep.
The study aims to investigate the effect and mechanism of PPARA on intramuscular fat (IMF) deposition,and to search for molecular markers related to IMF deposition in sheep.The experiment was conducted using Small-tail Han sheep (STH) and a hybrid F1 population between Suffolk sheep and Small-tail Han sheep(STH×SFK)as the research object.By measuring their meat quality,the histological structure and fat drop distribution of longissimus dorsi muscle of two sheep populations were compared by H&E staining and Oil Red O staining;qRT-PCR and WB techniques were used to detect the expression differences of PPARA mRNA and protein levels among different populations,and analyze the correlation between PPARA gene and IMF;Sanger sequencing technology was used to detect the PPARA gene SNP site to evaluate the possibility of using it as a genetic marker related to IMF deposition in sheep.The results indicated that the shear force and water loss rate of STH were significantly higher than those of STH×SFK,but pH was extremely significantly lower than that of STH×SFK,marble score of STH was lower than that of STH×SFK population;histological staining showed that STH muscle fibers were thick and tightly arranged,with IMF concentrated in the muscle fiber gaps,and muscle fibers were thin and loose in structure in STH×SFK,and IMF was evenly and widely distributed among muscle cell and muscle fiber spaces,with a higher content.The expression levels of PPARA mRNA in the STH were significantly higher than those in the STH×SFK population,and PPARA protein expressions of the STH were extremely significantly higher than those of the STH×SFK population.Correlation analysis showed that the expression level of PPARA gene was extremely significantly negatively correlated with sheep fat droplet area ratio and pH,extremely significantly positively correlated with shear force and water loss rate,and weakly correlated with marble score.Sanger sequencing results showed that base mutations T49885C,T50007C,G50013A,G50835A,G50942A and G51154C occurred in the second intron of PPARA gene in two sheep populations,but no C49885T mutation was detected in the STH population.The analysis of SNP genetic diversity showed that the homozygosity of SNP loci in the population was greater than the heterozygosity,and they were in a medium to low degree of polymorphism in the population;except for the G50835A mutation,all other SNPs were in line with the Hardy-Weinberg equilibrium,and with a stable genetic basis.It was believed that PPARA gene had an important impact on IMF deposition in sheep,and could be used as a potential molecular genetic marker for IMF trait selection in sheep.
The aim of this study was to investigate the effect of bisphosphoglycerate mutase(BPGM)on the growth and development of chicken myogenic cells.The transcriptomics and proteomics data was jointly analyzed in the preliminary stage of the study to identify the differentially expressed genes that might affect the growth and development,and investigated their effects on the proliferation,apoptosis and expression levels of related marker genes of myogenic cells.Inosinic acid(IMP)was detected by ELISA and adenosine triphosphate(ATP)was detected by colorimetric assay,which revealed the important roles of the key candidate genes on muscle development and meat flavor of Jingyuan chickens.The results showed that a total of 139 differentially expressed genes/proteins were identified between the breast muscle and leg muscle of Jingyuan chickens,including 75 differentially expressed genes/proteins that were up-regulated in both the transcriptome and proteome,45 differentially expressed genes/proteins that were down-regulated in both the transcriptome and proteome,13 differentially expressed genes/proteins that were down-regulated in the transcriptome,and 6 differentially expressed genes/proteins that were up-regulated in the proteome and down-regulated in the transcriptome.7 differentially expressed genes were screened from the glycolysis/glycolysis pathway,which were BPGM, phosphoglycerate kinase 1 gene(PGK1),glucose-6-phosphate isomerase gene(GPI),phosphoglycerate mutase 1 gene(PGAM1),lactate dehydrogenase A gene(LDHA),lactate dehydrogenase B gene(LDHB)and phosphoglycerate isomerase 1 gene(TPI1),and screened for the differentially expressed gene BPGM in combination with KEGG pathway enrichment analysis and protein network interactions analysis.BPGM was expressed in all tissues of Jingyuan hens,the highest expression was in pectoral muscle,and it showed an up-regulation trend during the differentiation of myoblasts.Primary myogenesis cells were isolated and cultured for cell transfection.Interfering with BPGM inhibited myoblast proliferation and promoted apoptosis;it also affected genes upstream and downstream of the glycolytic pathway,and inhibited the biosynthesis of IMP and ATP at the same time.In conclusion,down-regulation of BPGM inhibited the growth and development of myoblasts,as well as the biosynthesis of IMP and ATP.
The aim of this study was to analyze synaptosome associated protein 29 gene(SNAP29)in Cynoglossus semilaevis.The molecular biological characteristics,tissue distribution characteristics of SNAP29 gene and the temporal expression pattern after infection with Shewanella algae.The SNAP29 gene of C.semilaevis was amplified by RT-PCR,bioinformatics website and qRT-PCR,and its bioinformatics analysis and expression characteristics were studied.The results showed that the CDS region of SNAP29 gene was 789 bp in length,encoding 262 amino acids.The theoretical isoelectric point(pI)was 5.39,and the molecular weight was 29.717 81 ku.There were two typical SNARE domains.The secondary structure was mainly composed of α-helix and random coil,and the secondary structure and tertiary structure were basically the same.The results of multiple sequence alignment showed that the SNAP29 gene of C.semilaevis had the highest similarity with that of European perch.The SNAP29 gene was expressed in all tissues of healthy C.semilaevis,with the highest expression in gill tissue and the lowest expression in brain tissue.After artificial infection of C.semilaevis with S.algae,the expression of SNAP29 gene was mainly up-regulated in intestinal tissue,heart tissue and brain tissue,and down-regulated in liver,head kidney and spleen tissue.SNAP29 gene was expressed in heart,brain,liver,intestinal tract,spleen,gill,middle kidney and head kidney of healthy C.semilaevis.The SNAP29 gene was differentially expressed in the intestine,brain,heart,liver,head kidney and spleen of C.semilaevis infected with S.algae.The above results indicate that SNAP29 may be involved in the body's immune response process or pathophysiological process.
