To study the effect of different color bags on Marselan grape berry quality,and providing high quality raw material for vine culture.A complete randomized experiment design was used to analyze the effects of different color bags on berry quality(external quality and internal quality),peel quality and antioxidant activity.100 berries weight,reducing sugar,titrable acid and pH were increased in bags.The red bag increased the total phenols,total flavan-3-ols and total flavonoids,with 376.03,149.78,1 463.53 mg/kg,respectively,there were significantly differences for the total flavan-3-ols and CK,the total flavonoids and the CK.Blue and yellow bag treatments reduced the total phenols,total flavan-3-olsand total flavonoids.Bagging treatment significantly reduced total tannin content,the total tannin content of yellow bag was the lowest at 4.62 mg/g.Generally speaking,bagging treatment decreased DPPH free radical scavenging ability,FRAP(Reducing ability to Fe²⁺)and ABTS free radical scavenging ability,there was no significant difference for DPPH free radical scavenging ability and FRAP of red bag with CK,yellow bag significantly reduced FRAP,DPPH free radical scavenging ability and ABTS free radical scavenging ability.Correlation analysis showed that the soluble solids were positively correlated with reducing sugar and the sugar/acid ratio,the correlation coefficients were 0.98 and 0.95,respectively.In terms of peel quality,the total phenol was positively correlated with total flavan-3-ols and total flavonoids,the correlation coefficients were 0.95 and 0.98,there was a very significant positive correlation between total flavan-3-ols and total flavonoids,and the correlation coefficient was 0.98.For antioxidant activity,FRAP was very significant positively correlated with DPPH free radical scavenging ability and ABTS free radical scavenging ability free radical scavenging ability,the correlation coefficient were 0.74,0.86,respectively.Soluble solids,reducing sugars and sugar-acid ratio were significantly positively correlated with total anthocyanins,the correlation coefficients were 0.93,0.90 and 0.92,respectively.Principal component analysis was carried out on all the measured indicators,and two principal components with eigenvalues greater than 1 were extracted,the contribution rates of 78.31% and 10.09%,respectively,and cumulative contribution rates of 88.40%,which could basically represent most of the data of quality indicators.Red and blue bags were beneficial to berry quality,and the red bag was better,which can be used in Turpan region.

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.

To find out the effect of laying reflective film on fruit quality in facilities and the mechanism of promoting coloring,the changes of the appearance and anthocyanin content of Jingyan grape fruits were monitored with the film laid between rows,then transcriptome sequencing and differential expression gene analysis of the peels with different coloring were carried out using high-throughput sequencing and Illumina Novaseq 6000 sequencing platform.Finally,41 151 986—46 673 908 Clean reads were obtained from 12 samples,with Q30 was over 94% and Q20 was over 98%.Besides,the mapping ratio between Clean reads and grape reference genome was 92.69%—94.22%.The analysis results of differentially expressed genes (DEGS) showed that there were 3 768 DEGs in CK 0 d—CK 7 d,with 1 488 up-regulated and 2 280 down-regulated.And 5 129 in RF 0 d—RF 7 d after film mulching,with 2 048 up-regulated and 3 081 down-regulated.It showed that film mulching can increase the differential expression of genes.Compared with CK,934 DEGs were obtained after 7 days mulching,of which 441 genes were up-regulated and 493 were down-regulated.KEGG enrichment analysis showed that,a total of 179 DEGs were annotated on 64 signal pathways,which were mainly enriched in five metabolic pathways:metabolism,cellular process,environmental information processing,genetic information processing and organismal systems.Particularly,many genes vere differentially expressed in benzene propane biosynthesis,endoplasmic reticulum protein processing,plant pathogen interaction and plant hormone signaling pathway.Results showed some transcripts of PAL,F3'H and 3GT were up-expressed,and some auxin-related protein genes were down-regulated.For transcription factors,WRKY gene was mainly up-regulated,while bHLH,C2H2,NAC,MIKC-MADS,Dof were down-regulated.In conclusion,laying reflective film may improve the activity of phenylalanine ammonia-lyase(PAL)and the expression of related genes(F3'H,3GT),thus promoting anthocyanin synthesis.Meanwhile,transcriptional factors WRKY,bHLH,C2H2,MYB,NAC,MIKC-MADS and Dof may also play an important role in regulation.

In order to explore the effects of amino acid water-soluble fertilizer on grape yield and quality,the full-fruit Zaohebao grape was used as the experimental material,three treatments were set up:conventional fertilization (CK);increased application of organic fertilizer (OPT);organic fertilizer + amino acid water soluble fertilizer treatment (OPT+AA),the effects of exogenous amino acids on grape yield,growth,quality and soil nutrient content were studied.The results showed that compared with CK,grape yield decreased by 14.17% under OPT treatment and 7.50% under OPT+AA treatment,but were not significant.Compared with CK,partial productivity of fertilizer treated with OPT and OPT+AA increased significantly.Partial productivity of N,P₂O₅ and K₂O treated with OPT+AA increased significantly by 73.44%,85.01% and 85.01%,respectively.OPT+AA significantly increased total sugar by 8.09%,soluble solids by 1.32 percentage,sugar-acid ratio by 38.80%,and the titratable acid content by 0.16 percentage.The nutrient content of shoots was significantly increased by 24.73% in OPT+AA,total phosphorus content and total potassium content were significantly increased by 24.73%.In addition,OPT and OPT+AA treatments did not inhibit plant growth,but could effectively reduce the leaching of available nutrients and increase the nutrient content in the root domain.In summary,reducing and increasing the application of organic fertilizer had no significant effect on grape yield,but could reduce the leaching of available nutrients and significantly improve the partial productivity of fertilizer.The application of amino acid water-soluble fertilizer could promote nutrient absorption in plants,improving grape yield and fruit quality.

Grapes are often subjected to various abiotic stress during their growth and development,such as drought,salt,heat or chilling,among them,drought stress is one of the most important abiotic stresses inhibiting vegetative and reproductive growth of grape.Although grapes are more resistant to drought stress,about half of our country's main grape cultivation area belongs to arid and semi-arid climates,drought is one of the main factors restricting the development of our country's grape industry.To ensure the healthy development of our country's grape industry,the research on the effects of drought stress on grape,the formulation of rational irrigation system and the breeding of drought-resistant varieties have become the research hotspots in recent years.Due to the extensive effects of drought stress on grape,in older to cope with drought stress,grapes have evolved many regulatory mechanisms to balance the effects of drought stress in grape growth.Firstly,the water demand degree of grape in different growth period was analyzed according to the law of grape water consumption,and the effects of drought stress on main physical and chemical indexes were exhibited,including photosynthesis,osmotic regulation and reactive oxygen regulation.And then,the quality and yield of grape fruit were analyzed under drought stress,the effects of drought stress on grape fruit quality were reviewed.We give the prospect for how to use drought stress strategy to improve grape quality and how to further study the molecular mechanism of grape response to drought stress at last.

To clarify the relationship between BcPDR1 gene and MAPK pathway genes BcBMP1 and BcBMP3 of Botrytis cinerea.We analyzed the expression levels of BcBMP1 and BcBMP3 in mutants of BcPDR1 by Real-time PCR and the expression levels of BcPDR1 in RNAi mutants of BcBMP1 and BcBMP3.In the background of mutant ΔBcpdr1,strains ΔBcpdr1/BcBMP1-OE and ΔBcpdr1/BcBMP3-OE over-expressing BcBMP1 and BcBMP3 were constructed,and the phenotype and pathogenicity of the over-expressing strains were analyzed.It was found that the expression levels of BcBMP1 and BcBMP3 in BCt89 and ΔBcpdr1,gene mutants of BcPDR1,were significantly better than those of the wild-type strain and the revertant strain,and the expression levels of BcPDR1 in the RNAi mutants of BcBMP1 and BcBMP3 gene were obviously lower than those of the wild-type strain.The colony morphology,hyphal morphology,growth rate,sporulation,and pathogenicity of the over-expression isolates ΔBcpdr1/BcBMP1-OE and ΔBcpdr1/BcBMP3-OE were obviously different from ΔBcpdr1,and closer to the wild-type BC22.These results indicated that the BcPDR1 was closely related to the expression levels of BcBMP1 and BcBMP3 in B.cinerea.The BcPDR1 gene negatively regulates the expression of BcBMP1 and BcBMP3,while BcBMP1 and BcBMP3 genes positively regulate the expression of the BcPDR1.

In order to explore the mechanism of grape defense response to grape downy mildew,the leaves of Muscat and Moldova were inoculated with grape downy mildew in vitro and cultured in light incubator.The development of downy mildews was observed from macroscopic and microscopic aspect.In macroscopic,it was showed that white mildew layer appeared on the back of leaves of Muscat four days after inoculation,and the area of disease spots expanded with time,meanwhile chlorosis yellow spots appeared on the opposite position of the front side of leaves.However,there were only a few black spots on leaves of Moldova at the inoculation point.Under electron microscope,it was found that the spores of the pathogen invaded the stomata and colonized,and germinated new sporangium stalks,which released the spores and propagated on the leaves of Muscat.But the number of sporangia was small on the leaves of Moldova,and most of the stomata was closed,only with spores,hyphae and tissue fragments scattered around the stomata.Furthermore,transcriptome sequencing and differentially expressed genes were analyzed by Illumina high-throughput sequencing.A total of 664.86 Mb of high-quality reads were obtained by RNA-seq.GO enrichment analysis showed that differentially expressed genes were mainly concentrated in biological pathways(metabolic process,cellular process,response to stimulus and biological regulation category),cell components(cell, membrane, membrane part and organelle),molecular functions(catalytic activity and binding,transporter activity and transcription regulator activity).KEGG enrichment analysis showed that the differentially expressed genes were mainly enriched in the plant pathogen interaction pathway and MAPK signal transduction pathway.Nine genes that may be related to grape disease resistance were screened and verified by Real time PCR analysis,and the results were consistent with the transcriptome sequencing results.

To provide a theoretical basis for the high-quality production and precise fertilization of Merlot grapes,through the determination of the mineral element content of each growth and different tissue parts,and the nutritional diagnosis and analysis based on the quality index formed by the phenolic substances in the fruit.The comprehensive quality index of fruit phenolics,namely CI value,was calculated by Topsis analysis method.By comparing the correlation between content of various mineral elements in different growth stages/ tissues and CI value,the nutritional diagnostic factors are determined.In order to provide a theoretical basis for the high-quality production and precise fertilization of Merlot grapes,high-CI vineyards were classified by CND method.The results showed that there was a significant synergistic and antagonistic relationship among the mineral elements in the leaves,petioles and fruits of the Merlot grapes.VS_F_N,MS_F_P,VS_F_K,VS_L_Ca,FBS_F_Mg,VS_F_Fe,FBS_L_Mn,MS_F_Zn,FBS_P_Cu,VS_P_B,FBS_L_Mo were selected as plant nutrition diagnostic factors.The quality index inflection point value of high-CI vineyards was 0.735 5,of which 7 orchards met this condition,accounting for 14.58% of the overall sample.According to the mineral element content range of Merlot plant within high-CI vineyards,the appropriate values of nutritional diagnostic factors were:N(8.85—11.81)mg/g,P(1.98—4.26)mg/g,K(14.97—20.70)mg/g,Ca(35.57—68.83)mg/g,Mg(3.69—15.51)mg/g,Fe(70.96—103.26)mg/kg,Mn(166.20—277.67)mg/kg,Zn(10.71—20.27)mg/kg,Cu(9.54—14.90)mg/kg,B(11.44—17.07)mg/kg,Mo(0.69—1.60)mg/kg.The nutritional diagnosis of high-CI vineyards showed that the K,Ca,Mn,and Mg in Merlot plants were on the low side.The obtain the highest fruit quality index as the goal,the recommended fertilizer rates per hectare were N 62.25 kg,P₂O₅ 46.50 kg,K₂O 0.00 kg,CaO 56.25 kg,and MgO 46.50 kg.Ca and Mg fertilizers should be applied in small amounts and multiple times;we should strengthen the management of water and fertilizer in the orchard to increase the absorption of K element by the tree.Through foliar spraying,the lack of Mn elements can be appropriately supplemented.

The aim of this study was to investigate effects of wine quality in wine grapes Marselan on different rootstocks and provided theoretical reference for cultivation regulation.Wine grape Marselan grafted was used as experimental material in 2018,rootstocks were SO4,5BB,3309M and 101-14,respectively,own-rooted grape was used the control.The fruit was harvested when ripe in 2020 and 2021,bunch character and wine quality were measured.The results showed that density, bunch weight, average berry weight, berry length and berry width, maximum berry weight and No. of shriveling berry decreased,however,length and width and average berry weight of CK increased.Length and width,average berry weight and average seed number increased by grafting,the coefficient of variation of berry quality was small in 2 years.The berry weight of M/SO4,M/5BB and M/3309M mainly ranged from 1.01 to 1.25 g,M/101-14 mainly ranged from 0.76 to 1.00 g in 2020.The berry weight of M/5BB,M/3309M and M/101-14 mainly ranged from 0.76 to 1.00 g,M/SO4 mainly ranged from 1.01 to 1.25 g in 2021.Soluble solids decreased,however total acid mass concentration increased by grafting.Component and content of anthocyanins in peel were measured by LCMS/MS,the results showed that malvidins content was the highest,the second was delphinidins,pelargonidin was the least.Anthocyanins contents of M/5BB significantly increased by 26.62% compared with CK,anthocyanins contents of M/3309M and M/101-14 increased,but there was no difference with CK,anthocyanins contents of M/SO4 significantly decreased by 11.87% compared with CK.

In order to study the application method of humic acid on grapes and find out the optimal proportion of carbon such as humic acid instead of organic fertilizer,the plot experiment was carried out with simple chemical fertilizer fertilization mode as control(CK treatment)for two consecutive years.Based on farmers' fertilizer practice(FFP treatment),3 ratios of carbon such as humic acid instead of organic fertilizer were set:10% carbon substitution(T1 treatment),20% carbon substitution(T2 treatment)and 30% carbon substitution(T3 treatment).Their effects on grape yield,quality and soil nutrients were studied,and the optimal substitution ratio was screened by principal factor analysis.The results showed that the yield of T2 treatment was the highest for two consecutive years,but there was no significant difference between T2 and T3 treatment.Single cluster weight and single fruit weight in T1,T2,T3 treatments were significantly higher than those in CK treatment,but there was no significant difference between FFP treatment and T1,T2,T3 treatments;on the whole,the contents of soluble solids,soluble sugar,titratable acid and vitamin C in humic acid treatment instead of organic fertilizer were better than those in other treatments.Through correlation analysis,soil available potassium,organic matter and acid phosphatase were positively correlated with grape yield.Available phosphorus,available potassium,organic matter and acid phosphatase in soil were very significantly correlated with soluble solids,soluble sugar and vitamin C content of grape.Regulation of these soil indexes was the way to increase grape yield and quality by replacing organic fertilizer with humic acid;through principal factor analysis,there were 2 principal factors with eigenvalue greater than 1 in 2019 and 2020,the cumulative contribution rate of variance of 2 principal factors reached 88.864%-91.470%,and T2 treatment was the highest comprehensive score for two consecutive years.On the whole,carbon such as humic acid instead of 20% organic fertilizer was the optimal proportion.

The changes of minerals in leaves(L), flowers/fruits(F)and their correlations in the full-bloom stage(FBS), veraison stage(VS), and maturation stages(MS)of Merlot grapes in the Penglai producing area was analyzed, and which will provide the theoretical basis for the nutritional diagnosis and scientific fertilization of Merlot grapes. In 2018-2020, 16 Merlot grapes under fixed formula fertilization in the Penglai production area were selected as the research object. Each formula fertilizer selected 4 orchards, a total of 192 Merlot vineyards, and the leaves, flowers/fruits of different growth periods were compared. For differences in the content of mineral elements, correlation analysis was used to screen the main leaf nutrient factors that affect the mineral nutrient content of the fruit. The contents of L_Fe, B and F_N, P, K, Mg, Fe, and B content in each growth period showed the order of FBS>VS>MS, and there were significant differences in different growth periods. The contents of L_Mn, Cu and Cu content showed the order of VS>MS>FBS, and the differences were significant in each growth period. Except for N/Ca/Mg at FBS, Cu at MS, and Mn at FBS NS/MS, other leaf and flower/fruit mineral elements showed significant or extremely significant differences. The contents of N, Ca, and Mo in each growth period showed the order of L>F. The content of Fe, Mn, Zn, and B in each growth period showed as F>L. The content of P and K was expressed as F>L in both FBS and VS, and L>F in MS. The content of Mg and Cu in FBS was F>L, and in VS and MS, the content was L>F. There were similarities and specificities in the changes of mineral elements in the leaves and fruits of Merlot grapes, which might be caused by the different degrees of synergy or antagonism between the various mineral elements in the leaves and fruits. Correlation analysis showed that the content of various elements in leaves and fruits showed different degrees of correlation during different growth periods. P, K, Fe, Zn, Cu, B were indispensable elements in the development of Merlot grape flowers. The content of Ca, Mn, and Cu in the leaves and fruits of Merlot grapes increased significantly during the period of FBS and VS. During the MS of Merlot grapes, the N, P, K, Ca, Mg, and Mo contents in the leaves were significantly higher than those in the fruits, and the contents of Ca, N, and K were higher, indicating that the effects of Ca, N, K in the growth and development of the fruit were relatively high. The demand was higher than other mineral elements. In summary, the P, K, Ca, Mg, Fe, Zn, Cu, B, and Mo levels in the leaves before anthesis could be adjusted to regulate the content of P, K, Fe, Cu, and B in the flower;because the Ca in the leaves was not easy to effectively transport to the fruit for its use, so Ca should be directly applied to the young fruit in a targeted manner. During the VS, the content of Mn and Cu in fruits could be regulated by increasing the levels of Mn and Cu in leaves. During maturity, the contents of P, K, Zn, and Mn in the fruit could be regulated by increasing the levels of N, K, and Ca in leaves.

In order to provide reference for the selection of suitable scion-rootstock combinations in the production of wine grape in Ningxia.Chardonnay and Merlot were grafted with three rootstocks respectively to study the effects of rootstocks on the growth, fruit quality and yield of two kinds of grapes.The results showed that 5BB, 1103P and 110R promoted the growth of Chardonnay in different degrees, among which 1103P increased the trunk diameter, leaf area and SPAD value significantly, and the effect of increasing growth was the most obvious. 5BB significantly increased the reducing sugar and titratable acid content of Chardonnay fruit, while 1103P decreased the accumulation of reducing sugar and increased the content of titratable acid, and 110R had no significant effect on the sugar and acid content. 1103P and 110R significantly reduced the total phenol content of Chardonnay, while 5BB and 1103P increased the anthocyanin content significantly. There was no significant difference in yield per plant between the grafted and self-rooted Chardonnay. The growth of Merlot grafted onto 140R, 5BB and SO4 was slowed down evidently, among which 140R significantly reduced the trunk diameter, new shoot length and diameter, leaf area and SPAD value, and the effect of weakening growth was the most obvious. SO4 significantly decreased the reducing sugar content and increased titratable acid content of Merlot fruit, while 5BB had no significant effect on the reducing sugar content, but significantly increased the titratable acid content, and 140R had no significant effect on sugar and acid content. Three rootstocks improved the tannin and anthocyanin content of Merlot fruit, among which 140R showed a significant raise in tannin content, and the anthocyanin content of SO4 and 140R grafted was obviously higher than that of self-rooted. The rootstock 5BB significantly increased the yield per plant of Merlot.Among the three rootstocks, 5BB was the best one for Chardonnay;but for Merlot, the self-rooted seedling was the best, and the comprehensive performance of 5BB was better than the other two rootstocks in terms of stable growth, quality and yield.

For the problem of soil desertification caused by strong wind for long time and development of local grape industry in Yanhuai Basin,the influence of grasses on soil physical and chemical properties in the vineyards of Yanhuai Basin was determined,and the technology of planting grasses in vineyard was explored. Two kinds of grasses were selected for experiment and planted in 2017 according to the climatic conditions and characteristics of viticulture in Huailai County,and traditional cultivation as control. The physicochemical properties of sandy soil were analyzed after 3 years of planting. The results showed that:Compared with control,the nitrate content of 0-20 cm surface soil was increased significantly,and the available phosphorus,total nitrogen content of 0-80 cm soil layer was increased after planting Orychophragmus Violaceus (L.)O.E. Schulz for 3 years,while the physical properties of surface soil was improved.Compared with control,the organic matter and total nitrogen content of 0-20 cm surface soil was increased significantly after planting Carex lanceolata Boott. After 3 years of continuous cultivation,the contents of carbon and nitrogen in the aggregate of the surface soil were increased in vineyard.The influence of the two kinds of grasses on the aggregate structure of the sandy soil of the vineyard was different. The macroaggregate(>0.25 mm)content of surface soil was increased significantly after planting Carex lanceolata Boott and the microaggregate(<0.25 mm)content of surface soil was increased significantly after planting Orychophragmus Violaceus (L.)O.E. Schulz,so it was suggested that the surface soil should not be artificially ploughed after planting Orychophragmus Violaceus (L.)O.E. Schulz in order to prevent soil erosion in Yanhuai Basine. Soil physical and chemical properties could be improved and the effects on soil physical and chemical properties was different after planting grasses in vineyard,but the selection of suitable grass species was important in vineyard of Yanhuai Basin.

In order to study the effects of nitrogen application rate on the photosynthetic characteristics and assimilation product accumulation of Cabernet gernischet grapes at different growth stages, four urea application rates under drip irrigation conditions were set:150(N1), 300(N2), 450 (N3), 600 kg/ha(N4), 0 kg/ha was set to control(CK), the effects of different nitrogen nutrition on photosynthetic fluorescence parameters and carbon metabolism related indexes of grape leaves at growth period (5 days before flowering,20 days after flowering,50 days after flowering) were analyzed. The results showed that appropriate nitrogen application improved the chlorophyll content and RuBp carboxylase activity of leaves, enhanced the photosynthesis of vines, and increased the chlorophyll fluorescence of leaves which represented as the increase of effective photochemical efficiency (Fv'/Fm'), photochemical quenching coefficient (qP) and actual photochemical efficiency of PSⅡ(ΦPSⅡ) and the decrease of non-photochemical quenching coefficient (NPQ) in leaves. Compared with CK, Fv'/Fm', ΦPSⅡ, and qP under N2 treatment in DAF20 and DAF50 were significantly increased by 16.10%, 17.80%, and 21.49%, 21.13%, and 11.77%, 19.73%, respectively.Appropriate nitrogen application increased the activity of sucrose metabolism-related enzymes and soluble sugar content, but decreased starch content.And throughout the growth period, SPS, SS, and sucrose had the highest values under N3, glucose and fructose had the highest values under N2, while NI and AI had the highest values under N2, N2 and N1 respectively under DBF5, DAF20 and DAF50. Compared with CK, N2 treatment increased the content of soluble sugar, tannin, anthocyanin and the titratable acid content, promoted the fruit setting rate and the ratio of sugar to acid with yield of 14.50 t/ha.In conclusion, the application range of urea from 300 to 450 kg/ha enhanced the photosynthesis of leaves, promoted the synthesis of soluble sugars, thus providing sufficient nutrition for the growth and development of grape fruits.

The purpose of this study was to identify and analyze HXK gene family in grape. Based on the HXK(hexokinase) gene registration sequence of Arabidopsis thaliana and maize, four HXK genes were identified from the whole genome of Vitis vinifera, namd as VvHXK1, VvHXK2, VvHXK3 and VvHXK4. The bioinformatics analysis were performed on these four genes. The results of protein physicochemical properties showed that the number of amino acids encoded by the four genes was between 412 aa and 524 aa, of which VvHXK4 was a basic amino acid, and the other three were acidic amino acids. In addition, subcellular localization analysis indicated that all four VvHXK genes were localized in the cytoplasm. VvHXK4 was mainly localized in chloroplasts, and VvHXK2 might appear in mitochondria and nucleus. Meanwhile, the secondary structure prediction of the proteins found that the proteins encoded by the four VvHXK genes were mainly α-helix. The structural analysis of exons indicated that VvHXK1 contained 10 exons, and others contained 9 exons, showing that the VvHXK genes were relatively conserved; The initiation of cis-acting element analysis indicated that four VvHXK genes contained ABA response elements, MYB binding region and WRKY transcription factors, but no low temperature response component were found in VvHXK4. Phylogenetic tree analysis showed that the homology relationship between VvHXK1 and Arabidopsis AtHXK1 was the closest, indicating that it not only had the functions of regulating growth and development and affecting the growth of roots, but also had the effect of accelerating aging. The homology relationship between VvHXK2 and potato StHXK1 was recent, showing that it could increase the glucose phosphorylation activity in leaf extract or chloroplast. The homology relationship between VvHXK3 and potato StHXKRP1 was the closest, suggesting that VvHXK2 might be able to induce the expression of sugar in leaves. The homology relationship between VvHXK4 and spinach SoHXK1 was recent, which was important for studying the glucose metabolism of grapes. Fluorescence quantitative analysis showed that VvHXK2, VvHXK3 and VvHXK4 had the highest relative expression under the treatment of 20 g/L glucose, which was the most significant for glucose phosphorylation, and low expression under the treatment of fructose and sucrose. This is important for studying the sugar metabolism of grapes.

In order to define the characteristic aromatic components of each grape flavor type，we detected and analyzed the aromatic components of the berries from fifteen grape varieties (include 2 non-aromatic, 5 strawberry aromatic,8 muscat aromatic) using solid phase micro-extraction (SPME) coupled with gas chromatography-mass spectrometry (GC-MS).Results showed that the primary components of non-aromatic grape Moldova, Victoria is aldehydes. The relative content of 2-hexenal was the highest among aroma substances;The aroma substances of strawberry aromatic grape Shenfeng, Kyoho, Hupei No.2, Liaofeng, Shuofeng were mainly aldehydes, alcohols and esters; The aromatic substances of muscat aromatic grape were mainly aldehydes and terpenes. The main aldehydes are 2-hexenal and 1-hexanal.Terpenes were mainly linalool, dihydrolinalool, alpha-terpineol, nerol and geraniol. There were differences in aroma substances and substance contents among different grape varieties. This study can provide a theoretical basis for the research of grape aroma mechanism, breeding and promotion in the later stage.

In order to determine the methods of evaluating the environmental adaptability of different grape varieties in protected culture, it provides the theoretical basis for the selection of grape varieties. First, by measuring 22 kinds of grape cultivars photosynthetic physiological index and leaf physiological index which included light compensation point, apparent quantum efficiency, photosynthetic potential, total chlorophyll content, Chlorophyll a content, Chlorophyll b content, palisade tissue thickness, spongy tissue thickness, blade thickness, dry specific leaf weight,carbon dioxide compensation point, carboxylation efficiency, transpiration rate, dark respiration rate, water use efficiency and so on. Then using correlation analysis to find the significant correlation index with annual yield ability from these indices. Finally, based on the physiological indicators closely related to annual yield ability, using entropy method, Topsis comprehensive evaluation method and Topsis comprehensive evaluation method with entropy weight to analyze and compare the environmental adaptability of the grape facilities and taking annual yield ability as a verification indicator to screen out a quick and effective method for evaluating the environmental adaptability of the grapes in the facility. The result showed that：Centenial Seedless and Hongxiangfei grape varieties can be used as reference varieties for evaluating whether a certain grape variety is suitable for facility cultivation. Four parameters, such as light compensation point, chlorophyll a content, total chlorophyll content, and difference in photosynthetic potential inside and outside the facility, are significantly related to annual yield coefficient, and can be used as basic data for evaluating the environmental adaptability of the facility. Evaluation result of Topsis comprehensive evaluation method with entropy weight to the environmental adaptability of facilities grape is most similar with the result of annual yield ability evaluation, and Topsis comprehensive evaluation method is the second, and entropy method is the worst.

In order to provide a theoretical basis for the selection of suitable scion-stock combinations of wine grape in Xinjiang Manas River Basin, we studied the growth, photosynthetic property and fruit quality of different combinations. 2-year-old rootstock scion combinations of Marselan/SO4 and Marselan/5BB were selected as the treatments, and Marselan seedling was used as control. The growth, photosynthesis, chlorophyll fluorescence characteristics and fruit quality of Marselan were measured. The inter-segment thickness, inter-segment length, SPAD value and the fruit setting of Marselan/5BB were significantly higher than those of Marselan. The net photosynthetic rate(Pn), stomatal conductance(Gs)and PSⅡ actual photochemical efficiency(Φ_PSⅡ)of Marselan/5BB were the highest, while the intercellular CO₂ concentration(Ci)and the non-photochemical quenching coefficient(NPQ)was the lowest. Matheran/SO4 had the largest panicle quality, peel thickness and seed quantity, while grape skins had the smallest total tannin content. The total sugar content, total flavonoid content and total phenol content of grape skin in Marselan/5BB were the largest, but the seed weight was the smallest. According to the principal component analysis, the comprehensive scores were from high to low:Marselan/5BB > Marselan/SO4 > Marselan self-root seedlings. It suggested that the resistance of 5BB was better than that of SO4. Marselan/5BB was a suitable combination in Xinjiang Manas River Basin, possessing the great growth potential, high photosynthetic capacity and good fruit quality.

In order to explore the application effects of different organic fertilizers in vineyards and to provide a reliable basis for scientific fertilization of grape, the effects of three organic fertilizers(bio-organic fertilizer, sheep manure and pig manure)on root growth and soil nutrient status of grape were studied in a three-year-old organic vineyard of Xingcheng, Liaoning. The results showed that the application of bio-organic fertilizer increased the total root length and root surface area. The bio-organic fertilizer and sheep manure treatments could increase the total nitrogen content of roots(P<0.05), and the former increased more. Bio-organic fertilizer treatment significantly increased the contents of most mineral elements, especially in the harvest period except B element, and the peaks of other tested elements all appeared in the bio-organic fertilizer treatment group. Fertilization treatment significantly improved the soil organic matter content in the color change period and harvest period of grape(P<0.05), with the order of bio-organic fertilizer > sheep manure > pig manure > control, which increased by 70.72%, 53.67% and 52.31% respectively in color changing period, and by 51.61%, 36.14% and 34.84%(P<0.05)respectively in harvest period, compared to the control. The application of organic fertilizer could increase the content of alkali nitrogen and total nitrogen of soil at color-changing and harvesting stages, with the order of bio-organic fertilizer > sheep manure > pig manure > control, but only the organic fertilizer treatment had significant difference with the control in different periods(P<0.05). The soil available phosphorus contents in bio-organic fertilizer and sheep manure treatments was significantly higher than that of the control in the color change period and the harvest period, but there was no significant difference between the pig manure and the control(P<0.05). Fertilization treatment could lead to the increase of the content of most mineral elements in soil to a certain degree, while the B and Mo contents of soil in the color change period was lower than that of the control. The reason might be that the fertilization treatment enhanced the absorption of trace elements of grape and the trace elements in organic fertilizers released slowly. Compared with the control, the three organic fertilizers could improve the growth of grape roots, increase the content of mineral elements in roots, and increase the content of organic matter, total nitrogen, available phosphorus and mineral elements in soil. The effect of bio-organic fertilizer application was the best.

To explore the potential role of Resveratrol synthase(Rs) in fruit development of Vitis vinifera cv. Kyoho, the structure and function of Rs sequence were analyzed, and the spatio-temporal expression specificity at different developmental stages of fruit was identified. The bioinformatics analysis and qRT-PCR method were used to analyze the sequence of Rs gene in Kyoho grape. The structure and subcellular localization of Rs gene were predicted, and its expression in different tissues of fruit at different developing stages was analyzed. The full-length cDNA of Rs gene was 1 539 bp, with an open reading frame(ORF) of 1 179 bp, encoding 392 amino acids. The predicted molecular weight was 42.88 ku, and the theoretical pI was 6.09. The gene contained chalcone, stilbene synthase activity sites and complete stilbene synthase family characteristic sites. The protein interaction predicted that Rs interacts with OMT2.1, which catalyzes resveratrol biosynthesis of Pterotilbene. The results of subcellular prediction showed that the Rs gene mainly expressed in the cytoplasm and chloroplast. The promoter analysis showed that the expression of Rs gene might be regulated by light, MYB, fungi and hormones, presenting a certain tissue specificity. qRT-PCR analysis showed that the Rs gene of grape expressed at different stages of the peel and flesh, but the expression level was highest in the 25th day after anthesis. Combined with the role of Rs gene accumulation in resveratrol, it could be inferred that there might be a high accumulation of resveratrol in the fruit peel during the early period of fruit development. The Rs gene of Kyoho grape had a high conservation during evolution, and its expression might be regulated by environment, fungi and hormone. It had the interaction with OMT2.1 and had a certain specificity in different tissues and development periods.

In order to improve the effect of grape fruit coloring,the optimum concentration of ALA in solar greenhouse was discussed. In the enlargement and coloring period of the Flame seedless fruit,the use of 5-aminolevulinic acid 50,100,150 mg/L(ALA) spraying fruit,to investigate the effects of ALA treatment on color,quality and anthocyanin synthesis related enzyme activity of grape fruit appearance. The results showed that during the fruit ripening,the treatment of exogenous ALA could induce the accumulation of anthocyanin,promote the colouring of peel,and alleviate the degradation rate of chlorophyll and carotenoids; The content of soluble sugar in fruit increased significantly,and the content of titration decreased obviously. After treatment with ALA,the anthocyanin content of the peel was significantly correlated with PAL activity,there was no significant correlation with DFR activity,CHI activity was significantly related only after the treatment and the content of anthocyanin,but after treatment of ALA,PAL,CHI,DFR activity in pericarp than control had different degrees of improvement,by hypothesizing that the expression of ALA by upregulating the related regulation the factor to induce synthesis of anthocyanin accumulation,thereby improving fruit appearance and internal quality.

To solve the problem of grape root uplift and soil acidification,the influences in root growth and soil properties were studied by planting Lolium perenne L.and Medicago sativa L.in the inline of vineyard located at Xingcheng in Liaoning Province.The results showed that both Lp and Ms could significantly increase the root length and surface area in different stages and root levels (P< 0.05), and the amplification of which from high to low was Lp,Ms and control.The soil in vineyard by control management had an acidification trend,the pH distributed from 6.14 to 6.39.The pH ranged between 6.22 and 7.04 in Lp treatment,while that in Ms treatment was lower than Lp treatment.Compared with the control,the content of organic matter in Lp treatment significantly increased by 24.71%,48.07% and 44.44% (P< 0.05),while the Ms treatment increased by 7.87%,29.88% and 34.07%,respectively.In fruiting period,veraison and harvest time,the content of alkaline nitrogen in soil by Lp treatment increased by 40.40%,51.46% and 22.15% (P< 0.05) to the control,respectively. In fruiting period,veraison and harvest time,while those in the Ms treatment increased by 29.88%,28.03% and 5.42%,respectively.Compared with the control,both Lp and Ms treatments significantly increased the content of available phosphorus in soil (P< 0.05),which in Lp treatment was the highest,followed by the Ms treatment and the control.Both Lp and Ms treatments could cause the content of each mineral elements significantly increased at a certain time (P< 0.05).The overall effect of Lp treatment on the root length,root surface area of grape,and the content of organic matter,essential nutrients in soil was better than that of Ms treatment and control.

The extension of bio-control bacteria resources is significant to the development of bio-control of grape mycosis. To efficiently acquire bio-control bacteria again grape mycosis,the key components of the cell wall of fungal pathogen,chitin and β-1,3-gluncan,were successively used as the only carbon source. Through the screening method with substrate stress,target strains with both chitinase activity and β-1,3-glucanase activity were selectively enriched. Results suggested that this screening method effectively enriched the potential target strains. All of the 10 picked strains demonstrated chitinase activity and β-1,3-glucanase activity. Among them,the chitinase activity per biomass and β-1,3-glucanase activity per biomass of strain 1# reached 2.52,0.76 U/mL respectively,and those of strain 6# reached 1.45,3.99 U/mL respectively. Enzyme study proved that the enzymes of both strains were inducible. Under simulated natural conditions,the enzyme activities of the two strains reached the highest at 24 or 36 h. Through 16S rDNA sequencing,both of the two strains were identified as Pseudomonas sp.,which is a kind of endophytic bacteria with great potential for bio-control.

To investigate the expression of key enzymes affecting the composition of anthocyanins in the transformation stage of grape peel of Vitis amurensis. With the young leaves of Vitis amurensis as the test material, the genomic DNA of Vitis vinifera was extracted and the full length sequence of PAL (GenBank accession number:MH045991) was obtained by homologous cloning, and conducted a biological information analysis. The Q-PCR method was used to analyze the expression levels of 8 grapevine skins at different pigmentation stages. The results showed that the DNA sequence of the PAL gene was 1 763 bp in length and had a 1 671 bp open reading frame (ORF) encoding 556 amino acids. The molecular weight was 61.07 ku and the isoelectric point was 5.76, which was a stable protein. The whole polypeptide chain had a transmembrane helix structure with no signal peptide. Molecular phylogenetic tree analysis showed that the homology was higher with European and Asian grape species; The expression pattern of PAL gene in the 8 stages of the grape rind of P. villosa. The PAL gene was analyzed in the last 4 periods. With irregular expression levels, the PAL gene was hardly expressed during both the 50% color shift and 100% color shift. The regulation of PAL gene expression was regulated by many factors. It changed regularly in the early stage of growth and development of Vitis vinifera. In the last 4 periods, the expression level changes irregularly. PAL gene was in 50% color rotation and 100% color rotation. Almost no expression, which may be affected by some kind of inhibition of PAL enzyme activity factors.

The stress response and its physiological and relative gene characteristics of grape under the weak light conditions remains unclearly due to the lacking of research. The plant cell membrane system and metabolic process is also affected by low-light environment. As studies on response to weak light stress could provide for prevention of weak light adversity. Low light stress on one year Yinhong grape as the tested material with pot experiment, which was used to examine their growth changes, protective enzymes, osmotic adjusting materials, gene expression of MYC2 and TRX used Q-PCR under the different levels. Ranging from shading intensity of 0 to 85%(0, 25%, 40%, 70%, 85%).The results showed as follows:the growth and enzyme activities indicated that they all good at shading of T1, while the plant growth was gradually constrained with the enhanced weak light stress, SOD, POD, CAT, soluble protein activity, MYC2 and TRX gene expression increased firstly and then decreased gradually. At the same time, the activity of PRO and MDA was positively related with the degree of shade. The physiological indexes under higher intensity of shading (85%) were decreased significantly, which indicating that severe stress could cause irreversible damage by mediating itself and not adapt the weak light stress by regulating the cell protective enzymes and synthesizing the related genes. The conservation, the breeding of weak light varieties of grape was also discussed on the basis of the research results, it was also believed that more genes related to the weak light response need to be verified in the future to confirm whether they were induced by weak light.

To analyzed the relationship between BcKMO gene and MAPK signaling pathway in Botrytis cinerea,the specific inhibitor of MAPK signaling pathways U0126 was used to detect the sensitivity of the BcKMO gene mutants. It was found that the sensitivity of the BCG183 mutant to U0126 was remarkably weak compared to that of the BC22 and BCG183 /BcKMO. Expression patterns of BcKMO and MAPK signaling pathway key genes bmp1 and bmp3 were detected. The expression levels of BcKMO and bmp3 were stronger in 7 d mycelia and sclerotia of BC22. The expression levels of BcKMO, bmp1,and bmp3 were stronger in BC22 cultured on medium with sucrose or fructose. Real-time PCR technology was used to detect the expression levels of bmp1 and bmp3 in the BcKMO gene mutants,and the BcKMO expression levels in RNAi mutant of bmp1 and bmp3. The expression level of bmp1 was significantly upregulated in the BCG183 mutant,the expression level of bmp3 was significantly downregulated in the BCG183 mutant. The expression level of BcKMO in the RNAi mutants of bmp1 gene was obviously higher than that of BC22; the expression level BcKMO gene in the RNAi mutants of bmp3 was obviously lower than that of BC22. These results indicated that BcKMO negatively regulated the expression of bmp1 and positively regulated the bmp3 expression, bmp1 negatively regulated the BcKMO gene expression,and bmp3 positively regulated the BcKMO gene expression. Therefore,the results provide a foundation for clarifying the molecular mechanism of the BcKMO gene in growth,development and pathogenicity in Botrytis cinerea.

To explore the effects of different light qualities on the expression of CO4 gene in grapevine plantlet in vitro. The full-length cDNA sequence of CO4 was cloned from Vitis riparia×V.labrusca Beta in vitro by RT-PCR. Bioinformatics analysis of the second level structure, and phylogenetic tree of protein. The result showed that the size of CO4 gene fragment was 663 bp, the open reading frame (ORF)of CO4 gene was 747 bp, including 248 amino acid coding regions. By use bioinformation analysis, CO4 of Beta was a hydrophobic, unstable protein with good lipid solubility. CO4 had a signal peptide, it can be inferred that it was a transmembrane protein. Phylogenetic tree indicated that CO4 of Beta (Vitis riparia×V.labrusca)had the highest evolutionary relationship with Eucalyptus grandis (Eucalyptus grandis Hill ex Maiden).qRT-PCR analysis showed that the CO4 expressed specifically and strongly in the treatment of red to blue light (control), which increased 5.39 fold in comparison to the control. The difference was not significant when exposed to the treatment of white to red light and red to white light. However the expression of CO4 was the lowest in the white to blue light treatment, which was 54% of the control. Fluorescence parameter analysis showed that the maximum photochemical efficiency (Fv/Fm), potential activity (Fv/Fo)and photochemical quenching coefficient (qP)were significantly higher than those of the control under red to blue light. The non photochemical quenching coefficient (NPQ)was the lowest under that treatment, but was the highest under red light treatment, it was the lowest under this treatment. The CO4 of Beta was more sensitive to light. The treatments of red to blue light could not only promote the expression of CO4, but also could improve the Beta Grapevine in vitro photosynthetic capacity. Thus, red to blue light played an important role in the process of morphogenesis of Beta Grapevine in vitro.

To study the drug resistance of SCCmec resistant Staphylococcus aureus and analyze pubic health in Sichuan,we collected 120 samples from Sichuan pig-farms to identify MRSA by PCR test from Sichuan. Then identified these 22 MRSA by drug-resistance testing and SCCmec typing.In these samples,60 strains of Staphylococcus aureus had been isolated from pig water and the hand of worker,including 22 MRSA,the detection rate was 18.3% (22/120). Susceptibility testing results showed that this 22 multi-drug resistant MRSA strains had significant multi-drug resistance,most were 4,6-resistant. Genotyping results showed that the predominant type of the Sichuan MRSA were mainly SCCmec Ⅳ a type,including 17 strains of SCCmec Ⅳ a type and 5 strains of SCCmec Ⅰ type.Last discuss the MRSA from swine have imperiled the health of worker and public health。

For the further promotion of vegetative growth and effective fertilization in grape seedlings,taking the one-year-old potted Jumeigui grape seedlings as tested materials,using ¹⁵N tracer technique to study the effects of different fertilization methods (foliar fertilization and soil fertilization) on ¹⁵N-urea absorption,distribution and utilization.The whole plants were dissected 3 times to leaves,shoots,trunk and roots in mature stage of new shoots.The results showed that fertilizer was beneficial to shoot and leaf growth,and foliar fertilization had remarkable influences on total nitrogen of grape leaves on September 30.The Ndff values of different organs under different fertilization methods were different at the mature stage of new shoots.Ndff values in all organs under foliar fertilization treatment were significantly higher than those under soil fertilization treatment on September 10.On September 30,Ndff values of shoots reached to 10.03% when daubing on the leaves,which was higher than that when applying in soil (5.61%).The distribution ratios of ¹⁵N in leaves under foliar fertilization treatment were higher than those under soil fertilization treatment in three different period,which showed that foliar fertilization was beneficial to the absorption of nitrogen in leaves.¹⁵N utilization rate of daubing on the leaves were significantly higher than those of applying in soil on August 20 and September 10.On September 30,there was no significant difference of distribution ratios of ¹⁵N in different organs between two fertilizer manners,as well as the ¹⁵N utilization rate,and the difference of Ndff values in different organs reduced between two fertilizer manners.The foliar fertilization is beneficial to raise the ¹⁵N utilization rate in grape seedlings,and promote the vegetable growth.

In order to study the effect of 6-BA on grape fruit organic acid accumulation,and the possible molecular mechanism of 6-BA regulating organic acid synthesis,a grape cultivar,Rizamat was selected to study the effect of 6-BA on fruit organic acid accumulation and related genes expression. The inflorescences and fruit clusters were treated with 6-BA at 5 days before blooming,3 and 10 days after anthesis respectively.The results showed that organic acid content of the mature grape fruit was closely related to treatment concentration of 6-BA. Tartaric acid and total organic acid content of the mature fruits that treated by 30 mg/L 6-BA was significantly lower than control. But it didn't play important roles in the content of malic acid,citric acid and oxalic acid. For 10,20 mg/L 6-BA treatment,all kinds of organic acid didn't show significant difference between treated and control mature fruits. Gene expression result showed that the expression of IDH, MDH and PEPC were mainly suppressed by 6-BA treatment,but it also promoted the expression of ME. Therefore,we speculated that the reason for inhibiting organic acid synthesis was that parts of the organic synthesis related genes expression were affected by 6-BA treatment.

To analyses the effects of water deficit in different stages on the variation tendency of soil biological characteristics (soil invertase,urease,soil microbial biomass carbon,soil organic carbon)and the fruit yield and quality of greenhouse grape under delayed cultivation,the experiment was carried out at Zhangye Irrigation Experiment Station in Gansu Province in 2013-2015.The results showed:water deficit treatment during germination and shoot growth stage could improve soil invertase activity.There was no difference in water deficit treatment in flowering period compared with CK treatment.The treatment of water deficit at the fruit expanding stage and the coloring maturity stage could inhibit the activity of soil invertase.The germination period was the most suitable period to improve the activity of soil invertase by water deficit.The treatment of water deficit at the germination stage and at the color maturity stage could inhibit the activity of soil urease.The water deficit treatment from the shoot growth stage to the fruit expanding stage could improve the soil urease activity in the corresponding growth stage.For the soil microbial entropy,the treatment of water deficit at the growth stage of new shoots had a significant effect on it.According to the results of correlation analysis and canonical correlation analysis,the soil invertase activity increased with the increased of fruit yield,while the content of soil microbial biomass carbon decreased significantly.The content of titratable acid increased significantly when soil urease activity increased.

Using homology cloning method,the full-length cDNA of galactinol synthase gene VvGolS2, VvGolS3 and VvGolS4 were cloned from Vitis vinifera cultivar Zuoyouhong tissue culture seedling.The analysis of VvGolS2, VvGolS3 and VvGolS4 sequencing indicated that three genes were 954,978 and 1 011 bp ranging in size,coded as 317,325,336 amino acids respectively with molecular weight were 36.65,36.97,38.12 kDa and the isoelectric point were 5.12,5.33,5.16 respectively.Three VvGolSs all possessed a conserved hydrophobic APXAA pentapeptide domain in the C-terminal region.Real-time PCR analysis showed that three VvGolSs were expressed different in all tested tissues,with the highest expression of VvGolS2 and VvGolS4 in leaves,and VvGolS3 was highly expressed in flowers and tendrils. Using tissue culture seedling,the relative expression levels of VvGolS2, VvGolS3 and VvGolS4 after different stresses or different stress singling factors treatment were checked.The VvGolS2 and VvGolS4 were highly induced by salt stress,while the VvGolS3 was only induced by low temperature.Moreover, VvGolS2 was significantly induced by stress-related signal molecules,such as abscisic acid (ABA),ethylene (ACC),and hydrogen peroxide (H₂O₂).The VvGolS4 was induced by ACC,H₂O₂ and H₂S,and the VvGolS3 was induced by ABA and ACC.Together,these results suggested that these three VvGolSs were involved in abiotic stresses resistance in grape with different functions.

The aim of this study is prokaryotic expression analysis of BcKMO gene from Botrytis cinerea and obtain the purified BcKMO protein.The BcKMO gene was amplified by RT-PCR technology using the cDNA of the Botrytis cinerea wild type BC22,cloned into the pMD19-T vector and sequenced.The results of sequencing showed that the BcKMO gene sequence was right.The pMD19-T-BcKMO and pGEX4T-1 plasmids were digested using restriction enzyme.The BcKMO gene segments were collected and cloned into the pGEX4T-1 vector.The results of restriction enzyme digestion and sequencing showed that the vector pGEX4T-1-BcKMO-GST was successfully constructed.The vector pGEX4T-1-BcKMO-GST was transformed into E.coli BL21 strain.The results of IPTG inducement indicated that the pGEX4T-1- BcKMO -GST was successfully expressed in E.coli BL21 strain,with the molecular weight 71 kDa.The optimal conditions of the prokaryotic expression of BcKMO were determined as 0.2 mmol/L IPTG treatment 12 h.Western Blot results showed that the GST antibody could specifically bound to purified pGEX4T-1-BcKMO-GST fusion protein,suggesting that the expression of BcKMO gene was successfully in vitro.

In order to provide high efficient viticultural management in Beijing alluvial plain,we studied the water consumption regularity of grapevine growing in such areas. The experimental vineyard was located in Pinggu District,Beijing. Rain shelter cultivation,soil mulching and drip irrigation were used during the growing season,the row and plant spacing was 3 m×2 m. By using a soil moisture sensor(Beijing Insentek Corporation) in the whole year,the soil moisture data of different depth(10-30,0-50,50-70,70-90,90-110 cm) of six-year-old Ruiduxiangyu(Vitis vinifera) are collected. The decrement of soil moisture during day time was considered as water consumption while during night time was considered as infiltration. Followed by this logic,the water consumption of different soil layers in each phenological period were calculated;Combined with monthly precipitation during almost 30 years in Beijing,we found out the regularity of grapevine water consumption. The results indicated that:From January to June,the amount of water consumption was quite close to precipitation;While during July to September that contributed 78% to the whole year's precipitation,the rainfall during these three months was 300 mm more than the water consumption. The water consumption during each phenological period were as follows:budburst was 12 mm,shoot rapid growing was 24 mm,blooming was 30 mm,fruit development was 72 mm,verison was 43 mm,fruit maturation and nutrient back-flowing period were the same,about 40 mm.The daily water consumption was about 1 mm from budburst to shoot rapid growing;It increased to 2.1 mm per day during blooming and 2.4 mm during fruit development;Then it decreased to 1.34 m during verison and 1.0 mm from fruit maturation and nutrient back-flowing period. During the first half of growing season,the proportion of water consumption of 10-50 cm was almost 70%;The proportion of water consumption of 50-70 cm was 8%-10% during budburst and shoot rapid growing,while maintained around 13%-19% from blooming to nutrient back-flowing period;The proportion of water consumption of 70-90 cm was 10%-14% from budburst to verison while increased to 18%-20% from fruit maturation to nutrient back-flowing period;The proportion of water consumption of 90-110 cm was only 7% from budburst to verison,while increased to above 20% from maturation to nutrient back-flowing period. In conclusion,daily water consumption changes dramatically during the first half of growing season,it remained relatively stable during the latter period;The highest daily water consumption occurred at fruit development. It presented logarithm relationship that the water consumption decreased as the depth of the soil increased. Besides,water consumption of grapevine root system showed temporal and spatial variation,shallow root system consumed much more during the former period of growing season while deep root system showed the opposite.

This paper aimed to study the effects of nitric oxide(NO) and low temperature treatment on storage quality of grape fruits. Using Jufeng grapes as the experiment material,the grape fruits were soaked with 5,15 and 30 μmol/L NO solution and then stored at 0,25℃. The physiological index including fruits firmness,vitamin C,soluble protein,soluble sugar,reactive oxygen(ROS),malonaldehyde(MDA)and proline were investigated. Compared with 5 and 30 μmol/L NO treatment,15 μmol/L NO kept the grape fruits firmness effectively and delayed the concentration of soluble protein,the content of soluble sugar and vitamin C,also inhibited the increase of reactive oxygen and MDA content and increased the content of proline. 15 μmol/L NO made the grape fruits flavour kept long time during postharvest storage. However low temperature can keep grape fruits hardness effectively,reduce the content of MDA and reactive oxygen,slower the process of fruits rot greatly.The experimental results showed that 15 μmol/L NO and 0℃ storage could keep grape nutritional quality and delay the lower of fruits rot effectively,and then maintain the fruits flavour and enhance the fruits resistance.

Toll-like receptor (TLR) family plays a key role in innate immunity and various inflammatory responses,for recognizing the bacterial pathogen and mediated the signal path of inflammation.To investigate the expressions of TLR2 in mammary glands of Small-tail han sheep were infected with Staphylococcus aureu, and to explore the mechanism of TLR2 in mastitis,the qRT-PCR and IHC technology was used to detect the expression of TLR2 mRNA and TLR2 protein in the mammary gland of control and experimental groups.The results showed that the TLR2 mRNAs and protein were expressed in all mammary glands of Small-tail han sheep in control and experimental groups.The levels of TLR2 mRNA and protein was lowest in control group,was highest at 48 h after Staphylococcus aureu infusion,significantly higher than that of control and 96 h group(P<0.05).After infected 96 h,the levels of TLR2 mRNA and protein was significantly reduced than that of 48 h group(P<0.05).The protein of TLR2 were located in the epithelial cell of mammary gland acinus,were expressed in epithelial cell of mammary gland acinus dropped and inflammatory cells as lymphocytes after effected Staphylococcus aureu.The results implied that the expression of TLR2 in the epithelial cell of mammary gland was up-regulated to recognize pathogenic bacteria,arouse the innate immunity.

To investigate the function of VvMSA, the full-length cDNA of VvMSA was cloned from Vitis vinifera cultivar Vidal Blanc tissue culture seedlings.Bioinformatic analysis and quantitative Real-time PCR was used to analyze its characteristics.The results showed that VvMSA amplified fragment size of 450 bp encoding 149 amino acids with molecular weight 16.703 kDa,isoelectric point 5.68.VvMSA was an unstable hydrophilic protein and contained a conserved domain ABA/WDS encoding by 65 amino acids.VvMSA and the known homolog LeASR1 in tomato belongs to different sub-families.Real-time PCR analysis showed that the VvMSA was expressed in all tested tissues with the highest expression in flower. VvMSA was induced by salt,drought and cold stress in different levels,and highest expression level was showed after 3 h salt treatment.In addition, VvMSA was also induced by nitric oxide(NO),hydrogen sulfide(H₂S),abscisicacid(ABA) gibberellin(GA₃),auxin(IAA)and ethylene(ET),while salicylic acid(SA)inhibited its expression.The expression pattern of VvMSA by salt treatment was similar with the expression pattern by ABA treatment.In total,these results suggested that the VvMSA was involved in salt resistance by ABA signaling.

To study the expression of grape stress tolerance gene (CAN70200.1),a 1 354 bp promoter fragment (named as P_CAN) upstream of the gene CAN70200.1 was isolated by using PCR technology.Promoter sequence was analyzed by the database of PlantCARE and PLACE.The result showed that the P_CAN sequence contained basic elements CAAT-box,TATA-box and some cis-acting elements that response to abiotic stresses,light and plant hormones.To verify the expression pattern of the promoter,the P_CAN fragment was fused with GUS reporter gene located on pCAMBIA1391Z to construct a plant expression vector p1391Z-CAN,followed by transformation into tobacco by Agrobacterium-meditated method.The expression activity of P_CAN promoter reached highest at 120 min after drought stress treatment or at 30-60 min under 4℃ cold treatment condition,indicated that the P_CAN promoter could express under the condition of treatments with cold and drought.