Changes of chlorogenic acid content and its synthesis-associated genes expression in Xuehua pear fruit during development

According to synthetic pathway of plant chlorogenic acid （CGA）, the expression patterns of genes encoding enzymes that are associated with CGA synthesis were studied in normally developed Xuehua pear fruit. The study demonstrated that CGA content in peel and flesh of Xuehua pear decreased as fruit development progressed, with a higher level in peel. The expression levels of PbPAL 1, PbPAL2, PbC3H, PbC4H, Pb4CL 1, Pb4CL2, Pb4CL6, PbHC T1 and PbHC T3 genes decreased in fruit, which was consistent with the pattern of variation in CGA content. That indicated that these genes might be key genes for influencing fruit CGA synthesis in Xuehua pear. However, Pb4CL7 gene expression profile is not consistent with variation of CGA content, hence, it may not be a key gene involved in CGA synthesis.

Effects of 1-methylcyclopropene and modified atmosphere packaging on fruit quality and superficial scald in Yali pears during storage

The Yali pear（Pyrus bretschneideri Rehd.） is susceptible to superficial scald during prolonged cold storage and at shelf life. This study investigated the effects of 1-methylcyclopropene（1-MCP） and modified atmosphere packaging（MAP） on changes of fruit quality and superficial scald during cold storage and at shelf life in Yali pear. Compared with MAP, the combination of MAP and 1-MCP（MAP＋1-MCP） treatment reduced the carbon dioxide and ethylene content inside the packaging bag. The 1-MCP, MAP, and MAP＋1-MCP treatments reduced the superficial scald index, malondialdehyde content, O2^-· production rate and relative conductivity and inhibited the accumulation of α-farnesene and conjugated trienes in the peel. 1-MCP and MAP＋1-MCP treatments maintained a higher phenolic content and enhanced the catalase and superoxide dismutase activities in the fruit, while reduced activities of lipoxygenase and polyphenol oxidase in the peel preceding the onset of superficial scald. Comprehensive analysis indicated that the MAP＋1-MCP treatment is the most effective method tested for improving the quality of Yali pears during cold storage and at shelf life.

Combined application of Trichoderma harzianum SH2303 and difenoconazole-propiconazolein controlling Southern corn leaf blight disease caused by Cochliobolus heterostrophus in maize

Southern corn leaf blight(SCLB)disease caused by Cochliobolus heterostrophus is one of the major threats to corn production worldwide.The synergistic application of low toxic chemical fungicide and biocontrol agents could improve biocontrol stability and efficiency against plant diseases,which ultimately reduce use of chemical fungicide.Trichoderma spp.,well-known biocontrol fungi have been used to control some foliar diseases.However,few works have been reported on synergistic application of chemical fungicide and Trichoderma against foliar diseases.This study was aimed to investigate the control effect on the synergistic application of Trichoderma harzianum SH2303 and difenoconazole-propiconazole(DP)against SCLB.Results showed that the synergistic application of DP and SH2303 reduced the leaf spot area compared to the control.The efficacy of synergistic application of DP+SH2303 against SCLB could last for 15–20 d in pot trial under the greenhouse condition.Under the natural field condition,maize treated with DP+DP and DP+SH2303 showed 60%control,which was higher than that of SH2303+DP(45%)and SH2303+SH2303(35%).All these treatments induced the synthesis of defense-related enzymes(phenylalanine ammonia lyase(PAL),catalase(CAT),and superoxide dismutase(SOD))and the defence-related gene expression of SA pathway(PR1).Taken together the in-vitro leaf test and field trial,the control of SCLB by synergistic application of DP+SH2303 was similar to that of DP+DP.Among synergistic application,the sequential application of DP+SH2303 showed better control than the sequential application of SH2303+DP.It was concluded that the synergistic application of chemical fungicide(DP)and biocontrol agent(T.harzianum SH2303)could be used to reduce the chemical fungicide and to reduce the SCLB diseases in maize,which provided alternative approach to realize an eco-friendly controlling of the foliar disease.

Cloning,Localization and Expression Analysis of ZmHsf-like Gene in Zea mays

Using homology cloning method, a heat shock transcription factor （Hsf） like gene, ZmHsf-like, was cloned from maize （Zea mays） leaves. Sequence analyses showed that the open reading frame （ORF） of the gene ZmHsf-like is 1 404 bp long, encoding 467 amino acids. The sequence of amino acids encoded by ZmHsf-like contains the most conserved and typical DNA-binding domain of Hsf family. By bombardment into onion epidermis, we ifrstly found that the ZmHsf-like was subcellular-located in nucleus. NucPred analysis revealed there is a classic NLS of KKRR peptide in protein. Real-time PCR showed that ZmHsf-like gene expressed in leaves, stems and roots of maize seedlings under normal growth conditions, and the highest expression level was in roots, lower in leaves and the lowest in stems. The ZmHsf-like gene expression could be up-regulated by heat shock, PEG, ABA, and H2O2 in different degrees, among which the heat shock and ABA worked more efifciently. Obvious differences of the peak value and its corresponding time point of ZmHsf-like gene expression were observed among treatments. Experiments with inhibitor further suggested that the up-regulation ZmHsf-like gene expression of heat shock was H2O2-dependent while the induction of ZmHsf-like with PEG did not depend on the existence of H2O2. These results pointed out that ZmHsf-like gene probably regulates responsive reactions to abiotic stresses especially heat shock and drought through different signal transduction pathways.

Molecular identification of ω-secalin gene expression activity in a wheat 1B/1R translocation cultivar

ω-Secalin was an important factor influencing processing quality of wheat 1BL/1RS translocations. On the basis of four ω-secalin gene sequences cloned from Lankao 906 （a wheat cultivar with 1BL/1RS translocation） with putative transcription activity, a pair of primers with suitable restriction endonucleases added at their 5＂ ends were designed to amplify the mature protein-coding regions of the four genes. After digestion with restriction endonucleases, the amplified products were ligated into the prokaryotic expression vector pET30a（＋）. The prokaryotically expressed recombinant proteins and gliadin isolated from the Lankao 906 seed were analyzed on the same acid polyacrylamide gel electrophoresis. All four prokaryotically expressed secalin bands had corresponding seed-expressed gliadin bands. The four corresponding gliadin bands were confirmed to be the expression products of the four ω-secalin genes by liquid chromatography tandem mass spectrometry （LC-MS/MS）. This conclusion was further confirmed because the four ω-secalin bands could be detected in all 14 wheat 1BL/1 RS translocation cultivars used in the study, although there was some interference for the detection of one ω-secalin band from nearby wheat gliadin bands. The sequence information of ω-secalin genes with expression activity will be helpful for improving the processing quality of wheat with 1BL/1RS translocations by using RNA interference method to silence the expression of the ω-secalin genes.

Effect of Low Temperature Treatment on Agrobacterium-mediated Transformation of Maize Seed Buds

[Objectives]This study was conducted to improve the genetic transformation rate of maize. [Methods]The seeds of maize ＂ Zheng 58＂ as the experimental material were germinated and treated under freezing temperature of-18 ℃ for 30 and 35 min. The EGFP gene was transformed into the growth points of these seed buds by Agrobacterium tumefaciens EHA105. The transformation effect was determined by fluorescence protein detection on transformed buds. [Results]After a certain period of treatment at-18 ℃ following germination and transformation of maize bud growth points with A. tumefaciens,although the survival rate of maize buds was reduced,the percentage of transformed cells significantly increased. [Conclusions]Low temperature treatment can improve the transformation efficiency of A. tumefaciens to the growth point of maize bud.

The Physiological Basis of Heterosis for Potassium Uptake of Hybrid Millet

Hybrid millet Zhang and its parental cultivars were studied for their potassium (K) uptake and accumulation characteristics and related physiological mechanisms. Hydroponic culture was performed with two K levels (i.e., high and low) set up. At high K level, hybrid millet showed heterobeltiosis in K accumulation and leaf K content, and it also had higher H+-ATPase activity, respiration rate, root oxidant activity and root K+ influx rate than its parental cultivars. All these lay the physiological foundation of heterosis for potassium uptake of hybrid millet. At low K level, the hybrid millet had a lower H+-ATPase activity and a higher K efflux rate than its parental cultivars, thus heterobeltiosis in K accumulation or leaf K content was not observed. Therefore, high level K fertilizer application is recommended for hybrid millet cultivation as it is favorable for hybrid millet to display heterosis in K uptake and K accumulation.

NO_3^-/NH_4^+ ratios affect plant growth, chlorophyll content,respiration rate, and morphological structure in Malus hupehensis seedlings

Malus hupehensis （Pamp.） Rehd. is a widely cultivated rootstock in China. We studied the effect of three NO3-/NH4＋ ratios （100/0, 50/50, and 0/100, molar basis） at total nitrogen （N） concentration of 8 mmoL L-1 in a nutrient solution on M. hupehensis seedlings. Plant biomass, NO3- and NH4＋concentrafion, chlorophyll con- tent, respiratory rate, and cellular structure were investi- gated. M. hupehensis seedlings at the NO3-/NH4＋ ratio of 50/50 had the highest level of fresh weight, dry weight, shoot length, and chlorophyll （a, b, and a ＋ b） content, but the lowest respiration rate in the leavesand roots. In addition, thickness and numbers of palisade and spongy tissue cells of the leaves were greater with this treatment than with other treatments. At the NO3-/NH4＋ ratio of 100/0, the leaves and roots had higher NO3- concentration and lower NH4＋ concentration. However, the opposite trend occurred at the NO3-/NH4＋ ratio of 0/100. Chloro- phyll （a, b, and a ＋ b） content was lowest at the NO3-/NH4＋ ratio of 100/0 than at the other ratios. At the NO3-/ NH4＋ ratio of 0/100, oxygen （02） consumption increased in the leaves and roots, and irregular epidermis and cortex cells were observed in the root apical meristematic and mature region. Our results indicated that the NO3-INH4＋ ratio at 50/50 was suitable for growth of M. hupehensis seedling to achieve the highest biomass production and efficiency.

Effects of Acetylacetone Solution Soaking on Agrobacterium - transformed Maize Seed Buds

[Objectives]The effect of acetosyringone seed soaking on the transformation of maize seed buds was analyzed,so as to improve the genetic transformation efficiency of maize and to provide technical support for transgenic breeding of maize.[Methods]The seeds of the"Zheng 58"maize inbred line were used as experimental materials.When the seeds were germinated,AS was added to the water at concentrations of 70,140,210,and 280μmol/L,respectively,and the seeds germinated without the addition of AS served as the CK.The Agrobacterium-mediated method was used to transform bud growth points of maize seeds,and green fluorescent protein detection was performed on the young shoots transformed with EGFP(enhanced green fluorescent protein)gene.The effect of soaking seeds with acetosyringone solution on the transformation of maize bud growth points by Agrobacterium was studied according to the detection results.[Results]Soaking seeds in acetosyringone solutions for germination had the effect of inhibiting the germination of maize seeds and inhibiting sprout elongation,and the higher the concentration of acetosyringone,the stronger the inhibition.When the concentration of acetosyringone solution was 280μmol/L,the germination rate of seeds was only 36.2%of the CK,while soaking seeds with 70-140μmol/L acetosyringone solution for germination could not only ensure a higher germination rate of maize seeds,but also significantly increased the transformation efficiency of maize bud growth points.When the seeds were soaked with 70μmol/L acetosyringone solution for germination,the positive rate of transformed maize buds was the highest,reaching 32.1%.[Conclusions]When maize bud growth points were used as the receptor of Agrobacterium transformation,soaking seeds with 70-140μmol/L acetosyringone for germination basically did not affect the germination of seeds,and was beneficial to the activation of Agrobacterium,thereby promoting the transformation.

Study on Differentiation of Growing Points in Corn Sprouts into Multiple Stems

[Objectives] This study was conducted to improve the efficiency of genetic transformation using growing points of corn sprouts. [Methods] A mixed liquid of plant hormones 6-BA and KT at a suitable concentration and ratio was added dropwise to the growing points of corn sprouts, followed by culture under appropriate conditions. [Results] Corn sprouts could be induced to form multiple plants, thereby improving the transformation efficiency of growing points in corn sprouts. [Conclusions] This study provides a breeding method using growing points of corn sprouts as transformation receptors for direct plantlet formation without tissue culture by the Agrobacterium transgenic technology.

GmSAP5,a soybean A20/AN1 domain-containing stress-associated protein gene activated by GmAREB3,increases drought stress resistance in soybean by mediating ABA signaling

Drought stress impairs crop growth and productivity.Stress-associated proteins(SAPs),a class of zinc finger proteins containing the A20/AN1 domain,function in various stress responses in plants.However,little is known about the function of SAPs in drought-stress responses in soybean,an oil and protein crop.We report that a GmSAP5 protein confers drought tolerance by increasing sensitivity to abscisic acid(ABA)and reducing stomatal aperture.Overexpression and RNA interference of GmSAP5in soybean hairy roots resulted in elevated resistance and sensitivity to drought stress,respectively.ABA and proline contents increased in GmSAP5-overexpressing plants under water-deficit conditions.Lower water loss rates and higher relative water contents were observed in GmSAP5-overexpressing plants,resulting in increased drought-stress resistance.A yeast one-hybrid assay and luciferase transient transcriptional activity assay showed that GmAREB3,an AREB/ABF transcription factor,could bind to the promoter of GmSAP5 and activate its expression.These results suggest that GmSAP5 acts downstream of GmAREB3 and improves drought-stress resistance by mediating ABA signaling.

Boosting wheat functional genomics via an indexed EMS mutant library of KN9204

A better understanding of wheat functional genomics can improve targeted breeding for better agronomic traits and environmental adaptation.However,the lack of gene-indexed mutants and the low transformation efficiency of wheat limit in-depth gene functional studies and genetic manipulation for breeding.In this study,we created a library for KN9204,a popular wheat variety in northern China,with a reference genome,transcriptome,and epigenome of different tissues,using ethyl methyl sulfonate(EMS)mutagenesis.This library contains a vast developmental diversity of critical tissues and transition stages.Exome capture sequencing of 2090 mutant lines using KN9204 genome-designed probes revealed that 98.79%of coding genes had mutations,and each line had an average of 1383 EMS-type SNPs.We identified new allelic variations for crucial agronomic trait-related genes such as Rht-D1,Q,TaTB1,and WFZP.We tested 100 lines with severemutations in 80 NAC transcription factors(TFs)under drought and salinity stress and identified 13 lines with altered sensitivity.Further analysis of three lines using transcriptome and chromatin accessibility data revealed hundreds of direct NAC targets with altered transcription patterns under salt or drought stress,including SNAC1,DREB2B,CML16,and ZFP182,factors known to respond to abiotic stress.Thus,we have generated and indexed a KN9204 EMS mutant library that can facilitate functional genomics research and offer resources for genetic manipulation of wheat.