Scientia Agricultura Sinica

Difference of Elements in Different Types of Seedling Powder and Its Grains of Barley Recombinant Inbred Lines

ZENG Ya-wen, WANG Lu-xiang, YANG Xiao-meng, YANG Jia-zhen, DU Juan, PU Xiao-ying, DU Li-juan, YANG Shu-ming, XIAO Ya, YANG Tao

Scientia Agricultura Sinica. 2016, 49(15): 2857-2866. doi:10.3864/j.issn.0578-1752.2016.15.001

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【Objective】Variety type and element content as well as genetic variation based on seedling powder and grain for barley recombinant inbred lines (RILs) provide not only the basic information of barley grain and seedling powder for developing functional food that is better for human health, but also the evidences about genetic variation of cultivated barley. 【Method】Varieties classification of 194 RILs from a cross Ziguangmangluoerleng (ZGMLEL) ? Schooner and their parents can be made according to the types of ear and awn color, glume size, palea and lemma of lateral spikelets, awn shape, grain color, spike density, naked or husked seven traits. The contents of 9 mineral elements (P, Zn, Fe, Mn, Mg, Ca, Cu, Na and K) in seedling powder and grains of 194 barley recombinant inbred lines (RILs) and their parents were determined by using ICP-AES. 【Result】The 194 RILs based on hybridization breeding between Violiglabrum and Erectum as well as their parents were divided into 13 variety types, including Nutans, Hypianthinum, Lanthonudum, Violiglabrum, Medicum, Nudum, Erectum, Violimedicum, Neogenes, Glabrinudidubium, Jachanum, Luchuanicum, and Violierectum. It is obvious in genotype difference of variety classification traits. There are significant differences of minerial element contents among variety types in grains (1.9 to 6.4 times) and lower difference among variety types in barley seedling powder (1.2 to 2.1 times). Differences of the highest times for variety types in grain element contents is in order of Cu (Glabrinudidubium/Violierectum=6.4)＞Mn(Violimedicum/Violierectum=3.8)＞Fe (Neogenes/Glabrinudidubium=2.9)＞K(Medicum/Violierectum=2.3)＞Zn(Violiglabrum/Jachanum=2.2)＞P(Violimedicum/Jachanum=2.1)=Ca (Medicum/Violierectum=2.1)＞Mg (Violimedicum/Jachanum=1.9)=Na (Violimedicum/Violierectum=1.9); Differences of the highest times for variety types in seedling element contents is in order of Na (Violierectum/Luchuanicum=2.1)＞Fe (Neogenes/Erectum=1.5)＞Ca (Jachanum/ Luchuanicum=1.4) = Cu (Violiglabrum/Hypianthinum=1.4)＞Zn (Lanthonudum/Violimedicum=1.3) = Mg (Nudum/Luchuanicum=1.3) = K (Violierectum/Hypianthinum=1.3)＞P (Luchuanicum/Glabrinudidubium=1.2) =Mn (Nudum/Erectum = 1.2), however, the difference of 6 element contents (P, Zn, Mn, Mg, Ca and K) among varieties in grains and seedling powder is significant. The ratio of 7 element contents of barley seedling powder versus grains is 1.1-51.4 times. Differences of the highest times for variety types in element contents for seedling vs grains ratio is in order of Na (Violierectum 51.4)＞Ca (Violierectum 19.9)＞K (Violierectum 15.4)＞Fe (Glabrinudidubium 8.8)＞Mg (Jachanum 3.4)＞Mn (Violierectum 3.2)＞Zn (Jachanum 2.9)＞P (Jachanum 2.7)＞Cu (Violierectum 2.1). There are 194 RILs lines and their parents can be divided into 4 classes based on 9 mineral element contents for barley grains or seedling powder or seedling powder / grains, and the difference between groups is significant but the difference within a group is insignificant. The most variant type of their progenies are similar in mineral element contents in grains or seedling powder of Ziguangmangluoerleng with the original Yunnan landraces. 【Conclusion】There is a significant difference between groups but lower difference among variety types in mineral element contents in grain and seedling powder. There is a significant difference in mineral element contents in grain but lower difference in seedling powder. These evidences support that mineral elements play an important role in the domestication of cultivated barley variety type.

Molecular Clone and Expression of GhDHN1 Gene in Cotton (Gossypium hirsutum L.)

WANG Jun-juan, MU Min, WANG Shuai, LU Xu-ke, CHEN Xiu-gui, WANG De-long, FAN Wei-li, YIN Zu-jun, GUO Li-xue, YE Wu-wei, YU Shu-xun

Scientia Agricultura Sinica. 2016, 49(15): 2867-2878. doi:10.3864/j.issn.0578-1752.2016.15.002

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【Objective】In order to explore functional genes related to low temperature stress tolerance of cotton, the characteristics of cotton dehydrin gene and its expression patterns responsed to low temperature stress in cotton were analyzed, thus providing a theoretical basis for the application of dehydrin gene in cotton chilling tolerant breeding. 【Method】 In this study, based on the upland cotton genome sequence, specific primers were designed by Primer 5 software and the dehydrin gene was cloned from the upland cotton variety Yu2067, named GhDHN1. Bioinformatics analysis was conducted to analyze the properties, amino acids content, functional domains and evolutionary relationships of the gene. Plant expression vector pBI121::GFP at XbaⅠand SmaⅠ restriction sites was constructed with double enzyme digestions and the transient expression vector pBI121-GhDHN1::GFP was constructed by In-Fusion connection technology. And the subcellular localization of GhDHN1 was studied by transient expression analysis of onion epidermal cells. Combined with the transcriptome sequencing data of chilling-resistant cotton variety Yu2067, real-time fluorescent quantitative PCR expression of leaves, stems, and roots in chilling-resistant Yu2067 and chilling-sensitive variety Hengmian No.3 under low temperature stress treatments (4℃, 24 h) at trefoil stage was performed to study the function of GhDHN1. The expression of the gene in the leaves of two different cold resistant varieties was compared. The dynamic expression of GhDHN1 gene in leaves and roots of Yu2067 was detected under 4℃ low temperature treatment. 【Result】 A cotton dehydrin gene was cloned and the sequencing analysis showed that the cDNA of the dehydrin gene was 726 bp, and the gene encoded 211 amino acids with a predicted molecular weight of about 23.79 kD and the isoelectric point was 5.04. Amino acid sequence analysis indicated that the GhDHN1 was rich in glutamic acid content (26.10%) and lysine amino acid content (19.40%) with a half-life of 30 hours. GhDHN1 was acidic and negatively charged, of which 60 percent were negatively charged residues. The second structure analysis showed that the alpha helix of GhDHN1 contained 116 amino acid residues, accounting for 54.98% of the protein composition and random coil contained 87 amino acid residues. GhDHN1 gene was located on Dt_chr9 chromosome of AD genome. GhDHN1 gene contained a 90 bp intron at 259-348 position of cDNA and two exons in lengths of 258 bp and 378 bp, respectively. SMART and CDD analysis showed that GhDHD1 includes two conserved lysine-rich K fragments, a conserved serine-rich S fragment and a dehydrin functional domain pfam00257 and therefore was sorted into K2S subfamily of dehydrins. Phylogenetic analysis showed that GhDHN1 of cotton had the closest relationship with DHN1 of cacao. The analysis of the transient expression in onion epidermal cells showed that GhDHN1 was mainly localized near the cell plasma membrane. Transcriptome analysis showed that GhDHN1 gene was up-regulated in leaves and roots of cotton after low temperature treatments at the trefoil stage. qRT-PCR analysis showed that GhDHN1 was up-regulated in leaves, stems and roots after low temperature. The expression fold in leaves was higher than that in stems and roots. There were two expression peaks in leaves under low temperature treatment for 4 h and 24 h, and there were also two peaks in roots of the low temperature treatment for 6 h and 12 h. The leaf expression of the gene in cold resistant varieties was 2.47 times the expression in the cold sensitive varieties. The results showed that GhDHN1 gene may be involved in the adaptability regulation of low temperature. 【Conclusion】 GhDHN1 belonged to the member of K2S subfamily of dehydrins. Phylogenetic analysis showed that GhDHN1 had the closest relationship with DHN1 of cacao. GhDHN1 gene was induced by low temperature stress. Significant expression difference of GhDHN1 between the cold-resistance variety and cold-sensitive variety was found, and the expression of GhDHN1 was positively correlated with cotton cold-resistance. The expression of GhDHN1 can be thus employed as a marker of cotton chilling resistance. At the same time GhDHN1 gene can also serve as an important candidate gene in cotton cultivation of cold-resistant materials.

Genome-Wide Expression Analysis of Glucosinolate Biosynthetic Genes in Arabidopsis Across Diverse Tissues and Stresses Induction

DU Hai, RAN Feng, LIU Jing, WEN Jing, MA Shan-shan, KE Yun-zhuo, SUN Li-ping, LI Jia-na

Scientia Agricultura Sinica. 2016, 49(15): 2879-2897. doi:10.3864/j.issn.0578-1752.2016.15.003

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【Objective】The aim of this study is to analyze the expression profiles of Arabidopsis glucosinolate biosynthetic genes in different tissues at different developmental stages and under various stress treatments, so as to facilitate the identification of the key genes involved in controlling glucosinolate biosynthesis and provide valuable information for exploring the metabolic mechanism of glucosinolate biosynthesis as well as their functions in plant defenses. 【Method】 The dynamic expression profiles of 82 glucosinolate biosynthetic structural genes and regulating genes across a wide range of developmental stages and biotic and abiotic stresses were examined, based on ten expression microarray data and two RNA-Seq data in PLEXdb and AtGenExpress database. The expression patterns of ten key structural genes and regulating genes involved in glucosinolate biosynthesis pathway were further confirmed by Real-time PCR method. The correlation of the expression patterns of glucosinolate biosynthesis related genes was analyzed by STRING v10 software. 【Result】 The expressions of candidate genes between vegetative and reproductive organs were significantly different, with the majority of the candidate genes have higher expression levels in vegetative organs and lower expression levels in reproductive organs, indicating glucosinolates were mainly synthesized in vegetative tissues. The aliphatic glucosinolate biosynthesis genes were preferential expressed in shoot, such as stems and leaves, while indolic glucosinolate related genes were highly expressed in both of root and shoot, such as roots, stems and leaves. Moreover, the expressions of genes in a given tissue at different developmental stages were generally distinct, implying a temporal and spatial expression pattern. Real-time PCR analyses revealed that the expression profiles of the ten representive glucosinolate biosynthetic genes kept consistent with the results of microarray data assay. Furthermore, the expressions of aliphatic and indolic glucosinolate biosynthesis genes were tend to be induced by many bio- and abio-stresses, whereas indolic glucosinolate biosynthesis related genes were obviously induced by bio- and abio-stresses, such as Pseudomonas syringae, Alternaria brassicicola, Myzus persicae, low temperature (4℃), salt and high temperature (38℃). In addition, co-expression analysis revealed that the expressions of regulating genes were highly correlated to those of metabolic genes in aliphatic or indolic glucosinolate biosynthesis pathway. 【Conclusion】 The glucosinolates biosynthesis genes were preferentially expressed in vegetative tissues, as compared to seed tissues. There is a transport system that is responsible for transporting glucosinolates that were synthesized in vegetative tissues to seed tissues. The expressions of indolic glucosinolate biosynthesis genes tend to be induced by bio- and abio-stresses, indicating indolic glucosinolates are more important for Arabidopsis defense.

Responses of Soil Water and Temperature to Previous Wheat Straw Treatments in Plastic Film Mulching Maize Field at Hexi Corridor

YIN Wen, CHEN Gui-ping, CHAI Qiang, ZHAO Cai, FENG Fu-xue, YU Ai-zhong, HU Fa-long, GUO Yao

Scientia Agricultura Sinica. 2016, 49(15): 2898-2908. doi:10.3864/j.issn.0578-1752.2016.15.004

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【Objective】 Soil water and temperature are key factors for determining crop growth and resource use efficiency, response of soil water and temperature of crops to previous straw usually plays an important role in establishing efficient cropping systems and optimizing tillage measures. 【Method】 A field experiment was carried out in typical oasis irrigation region in order to optimize soil water and temperature of rotated maize under different previous wheat straw treatments and tillage modes, including no tillage with 25 cm height of wheat straw standing (NTSS), no tillage with 25 cm height of wheat straw covering (NTS), tillage with 25 cm height of wheat straw was incorporated in the soil (TIS), and conventional tillage without straw retention (CT). 【Result】 The results showed that NTSS, NTS significantly increased soil water content by 5.0% to 7.8% in 0 to 110 cm soil layer from sowing to seedling stage, 4.4% to 5.4% from jointing to big flare stage, 4.8% to 7.1% from silking to flowering stage, but there was no significant difference between NTSS and NTS, and NTS increased by 4.7% at filling stage, compared with CT. In particular, the treatment on NTS had advantage on maintaining high soil water content at whole growth period of maize. NTSS and NTS decreased evapotranspiration of rotated maize under plastic film mulching before silking stage, but evapotranspiration was increased after silking stages, which effectively coordinated water demand contradiction of maize growth at early and late stage, the effect of NTS was the best. NTSS and NTS optimized soil temperature of rotated maize under plastic film mulching, and the effect of NTS was obvious. NTS had higher soil temperature by 0.76℃ at eight o’clock in 0 to 25 cm soil layers in 2010, but lower by 3.67 to 3.87℃ at fourteen o’clock and 1.19 to 1.30℃ at eighteen o’clock, in 2010 and 2012, which indicate that NTS had effects on preservating soil temperature at low temperature stage at the day and night, and decreasing soil temperature at the high temperature stage. Meanwhile, NTSS and NTS reduced soil accumulated temperature in rotated maize field, and the reduction of NTS was more significant, it reduced by 67.1 to 76.2℃ from sowing to jointing stage, 29.3 to 50.5℃ from jointing to silking stage, and 46.7 to 75.3℃ from silking to late-filling stage, compared with CT. However, according to the value of difference between air and soil temperatures, NTS had the effect of maintaining soil heat in low temperature season and reducing soil temperature in high temperature season, which is an important regulation mechanism on growth and development of maize through reducing the excessive influence resulted from the temperature change. The grain yield of maize was 11.3% to 17.5% higher in the three straw returning treatments than that in CT check, NTS exhibited the most significant effect on high yield, reached 13 470 and 13 274 kg·hm^-2 in two study years, which were higher than TIS by 5.6% to 9.0%. 【Conclusion】 The results show that NTS treatment can be recommended as the best previous wheat straw treatment for optimizing soil moisture and temperature of rotated maize at oasis irrigation areas.

Comparison of Photosynthetic Gas Exchange Parameters for Sugarcane Close Relative Species Erianthus fulvus

XU Chao-hua, LI Chun-jia, LU Xin, LIU Xin-long, LI Xu-juan, MAO Jun, LIN Xiu-qin, LIU Hong-bo, ZI Qiu-yan, CAI Qing

Scientia Agricultura Sinica. 2016, 49(15): 2909-2920. doi:10.3864/j.issn.0578-1752.2016.15.005

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【Objective】A study on photosynthesis was carried out to examine the variation in gas exchange parameters among Erianthus fulvus clones, aiming at screening germplasm with good photosynthetic capacity and benefiting breeding for varieties with promoted photosynthetic capacity. 【Method】 Li-6400 photosynthesis system was used for gas exchange measurements of 36 clones of Erianthus fulvus at elongating stage.【Result】The each average value of gas exchange parameters was 17.61 µmol·m^-2·s^-1(A), 0.15 mol·m^-2·s^-1(Gs), 163.49 µmol·mol^-1(Ci), 2.17 mmol·m^-2·s^-1(E), 1.38 MPa (VPDL), 124.43 µmol CO₂·mol^-1H₂O(TE), 8.12 µmol CO₂·mmol^-1H₂O(WUE) and the fold change was 2.28, 2.88, 2.29, 1.96, 1.60, 1.93, and 1.75, respectively. Coefficient of variation of gas exchange parameters ranged from 11.79% to 26.79%, in which stomatal conductance had the highest value (26.79%), followed by the net photosynthetic rate a CV value of 21.48%, water use efficiency exhibited the lowest CV with a value of 11.79%. Statistical analysis showed that there was a significant variation in each gas exchange parameter among the clones. Gas exchange parameter correlation analysis showed that net photosynthesis rate was positively correlated with transpiration efficiency, stomatal conductance, water use efficiency, and a negative correlation with leaf vapor pressure deficit. Stomatal conductance showed a positive correlation with internal CO₂ concentration, transpiration rate, and a negative correlation with leaf vapor pressure deficit and transpiration rate. Environmental factors correlation analysis showed that leaf vapor pressure deficit had a significant correlation with longitude, but a significant negative correlation with elevation and altitude. According to the altitude, they were divided into high altitude and low altitude and high altitude of clones showed a better photosynthetic capacity than low altitude. Two principal components were picked up whose cumulative variance proportion was up to 89.79%. Elements of photosynthesis, conductance, transpiration rate, and those of internal CO₂ concentration, water use efficiency, transpiration efficiency were dominating ones for the first and second principal component, respectively. The 36 clones could be divided into 5 groups by systemic clustering analysis. Accordingly, the groups could be characterized by different levels of photosynthetic capacity or water use efficiency. Five models of fine discrimination were established, with 5 parameters selected from all the ones measured. The discrimination accuracy of the models was up to 99.03%.【Conclusion】The clones in group IV were high in both photosynthetic capacity and water use efficiency and also had rich materials, thus are potential to be used as parents in breeding progress.

Effects of DTA-6 and S3307 on Physiological Regulation in Normal and Abscission Pods of Soybean

CUI Hong-qiu, FENG Nai-jie, SUN Fu-dong, LIU Chun-juan, HE Tian-ming, ZHAO Jing-jing, LIU Yang, GONG Shen, SHI Chen, ZHENG Dian-feng

Scientia Agricultura Sinica. 2016, 49(15): 2921-2931. doi:10.3864/j.issn.0578-1752.2016.15.006

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【Objective】The objective of this experiment is to study the effects of plant growth promoter and plant growth retardator on the difference of physiology in normal and abscission pods of soybean during the pods development in the semi-arid area, to discuss the physiological effects of plant growth regulator on reduction of the abscission of flowers and pods in soybean, and try to find the ways to increase soybean output. 【Method】 Diethyl aminoethyl hexanoate（DTA-6）, Uniconazole（S3307）and water were foliage sprayed at initial flowering (R1) stage of 3 varieties including Suinong 28 (SN28), Kenfeng 16 (KF16), and Hefeng 50 (HF50) in the Lindian county, Daqing city of Heilongjiang province in 2012 and 2013. The first time to take samples was 35 d (R5) after spraying, since then, the normal and abscission pods treated with CK and treatments were collected every seven days. The pod coats and seeds were separated, quick frozen for 30 min in the liquid nitrogen, and then stored in low temperature refrigerator(-40℃). Determination was carried out when all the samples were collected. The regulating differences of DTA-6 and S3307 between normal and abscission pods in oxygen free radical metabolism, related abscisic enzymes and soluble matter in soybean were determined and compared. 【Result】 The results demonstrated that during the pod development process, the malonaldehyde (MDA) content, superoxide dismutase (SOD) activity, peroxidase (POD) activity, soluble sugar and soluble protein of abscission pods were significantly higher than normal pods, the abscission cellulose (AC) activity, polygalacturonase (PG) activity in abscission pods were significantly lower than normal pods. DTA-6 and S3307 could regulate the physiology of normal and abscission pods in soybean. Although DTA-6 and S3307 showed different process in regulating effect, but both in the process had many similar results. The regulating effects of DTA-6 and S3307 showed as follows: DTA-6 and S3307 treatments decreased MDA content, improved SOD, POD activities in normal pods, and decreased AC and PG activities at pod-filling early stage in SN28 and KF16, but decreased at pod-filling late stage in HF50. The physiological difference of normal and abscission pods could change with the pods developmental process in soybean. The extents of increase and decrease were different during pods development process. The physiological difference was influenced by genetically controlled factors.【Conclusion】Compared with normal pods, the physiological indexes of abscission pods were improved, plasma membrane peroxidation was enhanced, balance of protective enzyme system was broken, soluble material were increased, related abscission enzymes activities reduced, and these were affected by environmental factors. The normal pods regulated by DTA-6 and S3307 showed a positive response to biological membrane damage, osmotic adjustment, protective enzyme system, related abscission enzymes activity reduced, especially in improving the physiological regulation and self-repairing ability.

Guidance: Biological Control of Diseases and Insect Pests is Valid Method to Ensure Vegetable Safe Producing

LEI Zhong-ren

Scientia Agricultura Sinica. 2016, 49(15): 2932-2934. doi:10.3864/j.issn.0578-1752.2016.15.007

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Isolation and Identification of Lipopeptide Antibiotics Produced by Bacillus amyloliquefaciens B1619 and the Inhibition of the Lipopeptide Antibiotics to Fusarium oxysporum f. sp. lycopersici

XIANG Ya-ping, ZHOU Hua-fei, LIU Yong-feng, CHEN Zhi-yi

Scientia Agricultura Sinica. 2016, 49(15): 2935-2944. doi:10.3864/j.issn.0578-1752.2016.15.008

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【Objective】 Bacillus amyloliquefaciens, biocontrol strain B1619, can effectively prevent and control the occurrence and damage of tomato fusarium wilt. In order to study and analyze theantifungal substancesproduced by strain B1619, the lipopeptide antibiotics were isolated from the supernatant of fermented broth.【Method】Three primers designed based on the known lipopeptide genes sfp, ituA and fenB were used to amplify the corresponding genes from the genome of B1619 strain. The crude lipopeptides extracts were extracted by acid precipitation and methanol. The supelclean^TM LC-18 column and PHASE HF-NH2 column were used to separate lipopeptide antibiotics. All the samples were analyzed by HPLC and MALDI-TOF-MS techniques. Inhibition rates of the three lipopeptide antibiotics against Fusarium oxysporum f. sp. lycopersici were tested by dual culture on plate. 【Result】The PCR products with three primer pairs were cloned and sequenced. The result showed that the sfp, ituA and fenB genes existed in the genome of B1619. The HPLC results showed that the peak time of 60% methanol eluent isolates were at 12,16.5 and 18 min, the secretion was 18.5 mg·L^-1, and the retention times were consistent with bacillomycin L standards. The peak time of 70% methanol eluent isolates were at 22, 34, 37, 51 and 53 min, the secretion was 5.1 mg·L^-1, and the retention times were consistent with fengycins standards. The peak time of 100% methanol eluent isolates were at 27, 37, 41, 51 and 53 min, the secretion was 74.3 mg·L^-1, and the retention times were consistent with surfactins standards. The further verification of MALDI-TOF-MS results showed that 60% methanol eluent isolates which m/z were 1 043.4, 1 057.5 and 1 071.4 Da, was C13-C15 bacillomycin L, 70% methanol eluent isolates which m/z were 1 463.9, 1 477.9, 1 491.9 and 1 505.9 Da, was C15-C17 fengycins, 100% methanol eluent isolates which m/z were 1 008.6, 1 022.7 and 1 036.7 Da, was C13-C15 surfactins. The mycostatic tests showed that the antifungal activities of 1 mg·mL^-1fengycins were higher than 1 mg·mL^-1bacillomycin L, and the 1 mg·mL^-1surfactins had the weakest effect onF. oxysporum. However, when the concentration of surfactins was increased to 3 and 6 mg·mL^-1, the antifungal activities didn’t change obviously. 【Conclusion】B1619 strain secreted the three kinds of lipopeptide antibiotics in which fengycins and bacillomycin L played an important role in the inhibitory effect on F. oxysporum, while the surfactins had no obvious effect on it.

Effects of Bioorganic Fertilizers with Compound Microbes on Cucumber and Root-Knot Nematode

MA Yu-qin, WEI Cai, MAO Zhen-chuan, YANG Yu-hong, FENG Dong-xin, XIE Bing-yan

Scientia Agricultura Sinica. 2016, 49(15): 2945-2954. doi:10.3864/j.issn.0578-1752.2016.15.009

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【Objective】 The objective of this study is to evaluate the effects of bioorganic fertilizers with compound microbes (Brevibacillus brevis, Paecilomyces lilacinus, Acremonium implicatum, Trichoderma hamatum) on cucumber growth, yield and the control efficiency against root-knot nematode disease, then screen the efficient combinations of microbes.【Method】Experiments included 10 treatments of bioorganic fertilizers with compound microbes (No.1 to 10), which were from different compounds of the four kinds of microbes B. brevis, P. lilacinus, A. implicatum, and T. hamatum, the bioorganic fertilizers and the 10% fosthiazate granules were as the fertilizer and chemical control, respectively, the treatment without any fertilizer and pesticide was as the negative control. Based on the experiments performed in greenhouse and field, the differences of 10 treatments were analyzed by measuring the root-knot nematode disease index, cucumber yield, plant height and shoot fresh weight, and then the effects of bioorganic fertilizers with compound microbes on cucumber production and control efficiency of root-knot nematode disease were evaluated【Result】In pot experiments, the control efficiency of fosthiazate treatment was up to 89.2%, which was significantly higher than other treatments and controls. Among the 10 treatments of bioorganic fertilizers with compound microbes (No.1 to 10), the control efficiency of root-knot nematode of No. 5, No. 6, No. 9 treatments were 52.5%, 52.5%, and 54.2%, respectively, and the control efficiency of these treatments were higher than other fertilizer treatments. In the greenhouse plot, there was no significant difference in control efficiency among No. 5 (56.8%), No. 6 (57.6%), No. 9 treatments (59.3%) and fosthiazate treatment (61.3%). In addition, there was a significant positive correlation in the root-knot nematodes control efficiency between pot experiments and greenhouse plot, and the control efficiency of multi-microorganisms were better than that of single microorganism usually. Compared with the negative control, all fertilizer treatments increased cucumber growth and yield significantly (p＜0.01), but there were some difference among bioorganic fertilizers with compound microbes (No.1 to 10). In field trials, the yield increase was in the range of 31.7%-74.9%, the best treatment was No. 6, and the increased range of which was 74.9%. In pot experiment and greenhouse plot, compared with the negative control, the control efficiency of No. 9 ranked the best in yield, and the increased range was 60.4% and 68.1%, followed by No. 9 treatment, the control efficiency of No.1 were 50.8% and 55.8%, respectively. No. 5, No. 6 and No. 9 treatments could not only control the root-knot nematode disease effectively, but also promoted cucumber growth and increased their production, among them No. 9 treatment was the most effective treatment. There was a correlation in control efficiency of root-knot nematode and production increasing between pot experiment and greenhouse plot, but the correlation level was low. 【Conclusion】The bioorganic fertilizers with multi-microorganisms (No. 5, No. 6 and No. 9) have better effects on cucumber growth, yield and the control efficiency of root-knot nematode disease, thus possessing an important utilization value and research potential in agriculture production.

Natural Enemy Diversity on Trapping Crops and Its Application for Control of Aphids in Greenhouse Cucumber

WU Sheng-yong, XU Li-rong, LI Ning, WANG Deng-jie, LEI Zhong-ren

Scientia Agricultura Sinica. 2016, 49(15): 2955-2964. doi:10.3864/j.issn.0578-1752.2016.15.010

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【Objective】Concern over protection and application of natural enemies for biological control of insect pests has increased considerably in recent years. The objective of this study is to assess the relationship between natural enemy diversity on multiple mixed trapping crops and its influence factors. Furthermore, the efficacy of banker plant system for control of the target pest aphids in cucumber greenhouse was evaluated. 【Method】Five species of trapping crops including maize, alfalfa, sunflower, wheat and soybean were mix-planted in the experiment stations of Langfang and Huanggang in 2012, 2103 and 2015, respectively. The species of natural enemies were investigated during the experimental period. The community parameters of natural enemies were calculated, and the relationship between community structure and its influence factors (plant species, sampling time and flowering phase) was analyzed by redundancy analysis. In 2013, the dominant natural enemy, i.e., ladybird beetle was collected from the trap cropping and stored in laboratory. In addition, the wheat aphid and wheat were selected as the alternative prey and banker plant of ladybird beetle with different densities, respectively, and the system for control of Aphis gossypii was introduced on greenhouse cucumber. The control efficacy among different treatments was compared.【Result】Analyses of community parameters of natural enemies, Shannon-Wiener diversity index (H’), Simpson’s diversity index (D) and Pielou’s evenness index (J) showed differences over the sampling time. Aphidius spp. and Propylaea japonica were determined as dominant natural enemies in two years. The abundances of both species accounted for over 10% in all natural species on the mixed trap cropping. Redundancy analysis results indicated that plant species, flowering phase and sampling time significantly affected the abundance of natural enemies, and in total explained 26.02% and 17.20% of the variance of the natural enemy community data in 2012 and 2015, respectively. Graph of RDA in 2012 and 2015 indicated that Orius spp. and Chrysoperla sinica displayed higher correlation with the flowering phase. When introduced the banker plant system with density of 200 ladybird beetles in greenhouse, the density of A. gossypii was significantly lower among the treatments, with a control efficacy of 69.4%. 【Conclusion】Attractiveness of natural enemies was integrated with the strategy of their enhanced abundance by means of artificial assistance. Different species of trapping crop plants were mix-planted, attracted multiple natural enemies groups. The dominant natural enemy attracted in the mixed plants was screened to develop the banker plant system, which was then introduced in greenhouse cucumber for control of aphids. The banker plant system with the highest density of ladybird beetle resulted in a certain reduction in aphids population. Therefore, it is practicable to integrated strategies involving the trapping, protection, screening and application of natural enemies for biological control of insect pests.

Effects of Different Control Strategies on the Structure of the Arthropod Community in the Cauliflower Field

ZENG Liang-bin, CHENG Yi, YAN Zhun, MA Jun, REN Shun-xiang, WEI Lin, XUE Zhao-dong

Scientia Agricultura Sinica. 2016, 49(15): 2965-2976. doi:10.3864/j.issn.0578-1752.2016.15.011

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【Objective】The objective of this study is to reveal the effects of different control measures on characteristics of the arthropod animal community in the cauliflower field and provide a theoretical basis for pest control as well as protection and utilization of natural enemies. 【Method】The test ground of this work was divided into biological pesticide control area, chemical pesticide control area and non-pesticide control area. Five random sampling points were picked out and the numbers of all plants and arthropods on the test ground were recorded. Glass bottles with washing powders and water were placed on the set point of the field. The number of arthropod species in the ground was investigated. Cauliflower arthropod community in plots with different control measures were analyzed according to results. 【Result】There were 87 arthropod species in Guangzhou’s cauliflower field, including 29 insect pest species, 54 predator species and 4 neutral insect species. The control measures could obviously affect the diversity of species and the amount of each species of the arthropod communities. In non pesticide control area, the numbers of predators, insect pests and neutral insects were higher than those of the two pesticide control areas. The relationship of arthropod specie numbers and individual numbers among different areas was: non pesticide control area＞the biological pesticide control area＞the chemical pesticide control area. The dominant concentration of plant herbivore functional groups in cauliflower was at the minimum level at seeding stage and reached the maximum level at maturity stage. The dominant concentration of predatory natural enemies in different pesticide control areas was also relatively low at seedling stage. The concentration of the natural enemies remained at a similar level through different growth stages of the cauliflower in the non pesticide control area. On the other hand, the dominant concentration of predatory natural enemies fluctuated through different growth stages in pesticide control area. Cauliflower arthropod animal community varied at different growth stages and areas with different control measures. During the whole growth period, the species with the dominance index larger than 0.1 were Bemisia tabaci, Myzus persicae, Plutella xylostella, Pieris rapae, Pirata procurvus, Pardosa pseudoannulata and Coleosoma octomaculatum. The indices of cauliflower arthropod community diversity varied in different control areas and at different growth stages. Community overall diversity index (H′) increased following the cauliflower growth and reached maximum at late stage. These results coincided with the development of the field community. The trend of diversity index with different control measures was: non pesticide control area＞biological pesticide control area＞chemical pesticide control area. The variation of evenness index among different growth stages and different control measures was small. The variation trend in diversity of pest sub-communities and overall community was the same. The diversity index reached the maximum level in the late-mid period of cauliflower growth stage. The relationship among diversity indices with different control measures were: non pesticide control area＞biological pesticide control area＞chemical pesticide control area, which reached the minimum level at seedling stage of cauliflower in chemical pesticide control area. The evenness at cauliflower seedling stage was the maximum, and then decreased with the growth of cauliflower. In each growth period, the diversity of predator sub-community with different treatments remained relatively stable. The investigation results of the arthropod in the testing ground showed that the killing of the ground arthropod was larger in the two treatment areas. At the same time, it was possible to have a certain effect on the colonization of natural enemies at the early stage of the predatory natural enemies, according to the number of ground surface arthropod and the number of predatory natural enemies. Correlation of spiders with plant herbivore，neutral insects and predatory insects was significant. Correlation between predatory insects and neutral insects was also significant. These results indicated that between these functional groups in the number of compatibility or follow the effect. The correlation between predatory insects and insect herbivores was non-significant. 【Conclusion】The control measures could effectively reduce the population of pests, but also reduced the number of natural enemies. Neutral insects on ground surface would be helpful to early colonized predators. Chemical control could severely affect arthropod community of the cauliflower field, while the biological pesticide control has less influence on arthropod communities compared with the chemical pesticides.

Influence of Subculture on Virulence to Frankliniella occidentalis and Conidial Production of the Entomopathogenic Fungus Beauveria bassiana

ZHANG Hui, WU Sheng-yong, LI Juan, ZHANG Lu-lu, ZHANG Lin-ya, LEI Zhong-ren

Scientia Agricultura Sinica. 2016, 49(15): 2977-2987. doi:10.3864/j.issn.0578-1752.2016.15.012

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【Objective】The objective of this study is to assess the capability of the Beauveria bassiana against Frankliniella occidentalisin the field, and explore the influence of subculture on virulence and conidial production of the B. bassiana, propose an effective method for preventing the retrogression during successive subculture, and provide a basis for the large-scale production technology of B. bassiana. 【Method】 The virulence of 28 fungal strains to adult F. occidentalis was evaluated by soaking method with B. bassiana concentration of 1×10⁷ conidia/mL in the laboratory. The adults were dipped for 5 s in the conidial suspension. Conidial production of the 4 high virulent strains was measured on corn meal agar (CMA) medium and sabouraud dextrose agar yeast (SDAY) medium, screening high conidial production strain and spore production medium by comparing conidial production of different strains on different media. Subsequently, strain DZDC-9 was subcultured on three media (CMA medium, CMA medium with cicada exuviae, and CMA medium with F. occidentalis cadaver powder) to compare its virulence and conidial production over five subcultures. The relationship between conidial production and virulence was analyzed. 【Result】Among 28 strains of entomopathogenic fungi B. bassiana tested for virulence against F. occidentalis in laboratory bioassays, strains DZDC-9, GZGY-5, WLMQ1-8 and SZ-26 were found as the most potent. The corrected cumulative mortalities were over 90% after 5 days, and LT₅₀<3 d. Conidial production obtained from the CMA medium of each strain was higher than from SDAY medium, and strain DZDC-9 had the higher conidial production among the four strains. The results demonstrated that the virulence of strain DZDC-9 declined with successive subculture. However, when the strain was subcultured on the CMA medium with 0.5% cicada exuviae or 0.5% F. occidentalis cadaver powder as an additive, the virulence increased with successive subculture. Conidial production was not significantly different between the same generation subculture on CMA medium with cicada exuviae and CMA medium with F. occidentalis cadaver powder, but they were significantly greater than conidial production obtained from CMA medium. A positive correlation was found between sporulation and virulence of B. bassiana conidia under different media and culture conditions.【Conclusion】The potential of B. bassiana strain DZDC-9 for the control of F. occidentalis was studied, and proposed an effective method for stabilizing the virulence and sporulation of B. bassiana. Conidial production could be used as an indicator for virulence evaluation.

Responses of Soil Organic Carbon, Organic Nitrogen and Nitrogen Supply Capacity to Long-Term Nitrogen Fertilization Practices in Dryland Soil

WANG Hui, LIU Jin-shan, HUI Xiao-li, DAI Jian, WANG Zhao-hui

Scientia Agricultura Sinica. 2016, 49(15): 2988-2998. doi:10.3864/j.issn.0578-1752.2016.15.013

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【Objective】The purpose of this study is to reveal the response of soil organic carbon, organic nitrogen and soil N mineralization to different N application rates and their relationships, and then to evaluate soil N supply capacity in dryland soil. 【Method】A ten-year-old winter wheat (Triticum aestivum L.) field experiment with N application rates (0 (N0), 160 (N160) and 320 (N320) kg·hm^-2, plus 100 kg P₂O₅·hm^-2) in the experiment farm of Northwest A&F University, Yangling, Shaanxi Province was conducted to determine soil organic carbon (SOC), N (SON), microbial biomass C (MBC) and biomass N (MBN) and a laboratory incubation method was used to determine the capacity of soil nitrogen (N) mineralization. 【Result】Compared with N0 treatment, N fertilization treatments (N160 and N320) significantly increased SOC at 0-10, 10-20, 20-40 and 0-40 cm soil depths, and they had different effects in the stages of wheat pre-sowing and harvest; N160 and N320 treatments also significantly increased SON at 0-10 cm soil depth, and N320 treatment only increased SON at 0-40 cm soil depth; N fertilization treatments (N160 and N320) did not change MBN and MBC concentrations at 0-10 and 10-20 cm depths, while N320 treatment significantly increased MBN concentrations at 20-40 and 0-40 cm depths. In 0-10 cm soil depth, the amount of cumulative mineralized N and potentially mineralizable N (N₀) had a significant positive correlation with N fertilizer rates and SON, while N mineralization rate constant (k) had a negative correlation with them. In 10-20 cm soil depth, during the whole incubation process, the amount of cumulative mineralized N in different N treatments was significantly higher than that in N0 treatment, with the increment of 27.3% (N160) and 35.2% (N320), respectively, and there was a significant positive correlation between cumulative mineralized N and SON, SOC as well as N rates. The potentially mineralizable N (N₀) significantly increased with the increment of N fertilizer application rate, while the N mineralization rate constant (k) decreased. In 20-40 cm soil depth, the amount of cumulative mineralized N increased with the treatment of N320 and had a significant positive correlation with soil MBC and SON. 【Conclusion】Applying a reasonable rate of N fertilizer is one of the effective practices to improve the soil organic N and organic C concentrations in the dryland of Loess Plateau, and can also increase the soil potentially mineralizable N, reduce the N mineralization rate, thus improve soil N supply capacity.

Soil Amendment and Enrichment Efficiency of Agro-Forestry Models in Serious Earthquake Region of Northern Sichuan Basin

ZHAO Chang-ping, WANG Jing-yan, GONG Wei, YAN Si-yu, SHU Zheng-yue, CAI Yu

Scientia Agricultura Sinica. 2016, 49(15): 2999-3009. doi:10.3864/j.issn.0578-1752.2016.15.014

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【Objective】The effects of different agro-forestry models on soil amendment and enrichment efficiency were studied to select the optimum model in serious earthquake region of northern Sichuan basin.【Method】Soil physical properties, nutrient contents and enzyme activities were determined under 8 agro-forestry models (including Pyrus spp+canna edulis (LSJO), Pyrus spp+Vicia faba (LSHD), Pyrus spp+Brassica chinensis (LSYC), Pyrus spp+Ipomoea batatas (LSHS), Juglans regia+Canna edulis (HTJO), Juglans regia+Amorphophallus rivieri (HTMY), Eriobotrya japonica+Glycine max (PPDD), Eriobotrya japonica+ Ipomoea batatas (PPHS)) and farmland control (CK), and the membership function method was used to evaluate soil anti-erodibility, fertility, and amendment and enrichment efficiency of different models.【Result】Compared with the CK, soil non-capillary porosity, capillary porosity, total porosity, ventilation degree, natural water content, maximum water-holding capacity, capillary water-holding capacity, minimum water-holding capacity and drainage capacity of the 8 agro-forestry models were increased by 17.6%-161.8%, 11.6%-32.7%, 12.5%-45.2%, 17.9%-79.5%, 10.7%-35.4%, 13.7%-48.6%, 12.0%-33.1%, 16.4%-58.7% and 10.4%-25.3%, respectively; ＞0.25 mm soil aggregate (dry sieving), ＞0.25 mm water-stable aggregate (wet sieving) and mean weight diameter of water-stable aggregates were increased by 0.9%-7.2%, 5.6%-18.1% and 14.8%-138.7%, respectively; ratio of soil structure deterioration and index of unstable aggregate were reduced by 24.0%-51.4% and 17.1%-54.7%, respectively; ＜0.002 mm clay content, structure particle index, aggregation status, aggregation degree and physical stability index were increased by 15.1%-45.2%, 14.2%-28.9%, 69.3%-417.3%, 58.3%-256.6% and 3.5%-23.9%, respectively; ＜0.05 mm micro-aggregates content, dispersion rate, erosion coefficient and eroded index were reduced by 5.4%-33.7%, 8.4%-44.1%, 18.0%-49.8% and 19.1%-75.1%, respectively; organic C, total N and hydrolysis N content were increased by 7.1%-46.7%, 4.3%-30.9% and 18.8%-57.5%, respectively; available P and available K content were reduced by 1.7%-29.7% and 20.8%-53.4%, respectively; invertase, phosphatase and urease activity were increased by 17.3%-60.0%, 34.7%-149.2% and 21.0%-102.8%, respectively. The comprehensive value of soil anti-erodibility (CVSA) and amendment and enrichment efficiency (CVAE) in all agro-forestry models were significantly higher than that in CK. The comprehensive value of soil fertility (CVSF) in all agro-forestry models except for LSHS and PPHS were significantly higher than that in CK. Planting canna edulis, canna edulis and Glycine max under Pyrus spp plantation, Juglans regia plantation and Eriobotrya japonica plantation respectively, could gain higher CVSA, CVSF and CVAE. The CVSA positively and significantly correlated with CVSF (P＜0.05). 【Conclusion】Planting canna edulis, Vicia faba, Brassica chinensis in Pyrus spp plantation, planting canna edulis and Amorphophallus rivieri in Juglans regia plantation and planting Glycine max in Eriobotrya japonica plantation have significant soil amendment and enrichment efficiency, and are important to improve soil anti-erodibility and soil fertility in serious earthquake region of northern Sichuan basin. The improvement of soil anti-erodibility and soil fertility has a synergistic effect.

Effect of 24-epibrassinolide Treatment on Grapevine Leaf Against Plasmopara viticola

LIU Qing, LUAN Xue-tao, XU Shi-yan,MENG Ying, GAO Jiang-man, XI Zhu-mei

Scientia Agricultura Sinica. 2016, 49(15): 3010-3018. doi:10.3864/j.issn.0578-1752.2016.15.015

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【Objective】 The study researched the effect of24-epibrassinolide (EBR) treatment on the resistance in grapevine leaf against Plasmopara viticolaand the mechanism of the induced resistance. 【Method】 Cabernet Sauvignon (Vitis vinifera L.) grapevine leaves were used for experimental materials. At the early development of P. viticola invading grapevine leaf, the effect of different concentration of exogenous EBR (0.1, 0.5 and 1.0 mg·L^-1 EBR) on the disease incidence and severity of downy mildew of grapevine leaves, linear length of colonies per infection, the development of P. viticola sporangiophores while this pathogen invading the grapevine leaf, the number of spores per stoma, the stomatal aperture and the content of endogenous hormone and the relation between hormone content and stomatal aperture were investigated. 【Result】 EBR-treated leaves had a significantly lower stomatal aperture at 0.5 h after inoculation. Both on 1 d and 2 d, a significantly lower linear length of P. viticola colonies per infection was observed in all EBR treatments. The spread of mycelium almost covered most of the infection area in control while 0.5 and 1.0 mg·L^-1 EBR treated leaf resulted in a markedly restricted number of spores per stoma and growth of P. viticola after 3 days of inoculation. After 4 days of inoculation, EBR treatment significantly controlled the development ofP. viticola sporangiophores, disease incidence and disease severity, and 0.5 mg·L^-1 and 1.0 mg·L^-1 EBR treatment resulted in higher resistance. The leaves treated with 0.5 mg·L^-1 and 1.0 mg·L^-1 had a lower disease incidence and severity of downy mildew, while disease incidence was decreased by 51.4% and 45.0%，and disease severity drop by 71.2% and 62.9%, and there is no significance between the two treatment. There was a significant difference in ABA, JA and SA contents in grape leaves between CK and 0.5 mg·L^-1 EBR treatment. Stomatal aperture has significantly positive correlation with SA content while ABA content does with JA content. 【Conclusion】 The increased resistance against P. viticola invasion was possibly related with the suppression of pathogen development and the stomata closure which plant hormone crosstalk involved in.

Preparation and Antimicrobial Activity of Cottonseed Protein Hydrolysate

HAN Xiao-yan, BAO Yu-ming, XIN Feng-jiao, KONG Zhi-qiang, Christophe Blecker, DAI Xiao-feng

Scientia Agricultura Sinica. 2016, 49(15): 3019-3029. doi:10.3864/j.issn.0578-1752.2016.15.016

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【Objective】 In order to provide a theoretical basis and technical support for efficient utilization of cottonseed protein, two methods were used to prepare cottonseed protein, then the isolated proteins were compared and digested in vitro by pepsin and trypsin, lastly the hydrolysates were detected and selected in antibacterial activity. 【Method】 In this paper, cottonseed protein isolate was prepared by the traditional method which is alkali extraction and acid precipitation and Osborne fractionation method. The protein content and molecular weight of the obtained proteins were determined by the Kjeldahl method and Tricine-SDS-PAGE respectively, microstructure of the proteins were examined by scanning electron microscopy. Amino acid composition of cottonseed proteins were analyzed by automatic amino acid analyzer (L-8900). And then proteins were digested by pepsin and trypsin in vitro in the process of digestination, the degree of hydrolysis (DH) of cottonseed protein isolate (CPI) and cottonseed globulin (CPG) hydrolysates were determined by ortho-phthalaldehyde (OPA), and then kanamycin (Kan) and protein solution without hydrolysis were used as control to determine the antibacterial activity of protein hydrolysates with Staphylococcus aureus and Escherichia coli bacteria. Meanwhile, the molecular weight distribution of the hydrolysis products were detected by high performance liquid chromatography (HPLC). 【Result】The extraction rate of CPI by the method of alkali extraction and acid precipitation was (70.52±2.40)% and its protein content reached (89.53±0.66)%. While the albumin, globulin, prolamin and glutelin were classified by Osborne fractionation method, and compared with the other three proteins, cottonseed globulin got the highest protein content that was (82.57±1.02)%. And there was no significant difference (P＞0.05) in protein extraction rate between the two preparation methods for the proteins extraction rate of Osborne fractionation method was (67.55±1.16)%. The Tricine-SDS-PAGE pattern showed that the subunits composition of CPG was more closely to CPI, compared to the other obtained proteins by Osborne fractionation method. The microstructures of the obtained proteins were different from each other. The structure of CPI showed a neat honeycomb structure, which was similar to glutenins’ microstructure, while the CPG showed a uneven granulated one. Amino acid composition analysis indicated that the amino acids of CPG were well maintained by Osborne method, compared to CPI. Cottonseed proteins were hydrolyzed by pepsin and trypsin simulated the digestive system in vitro. The hydrolysis degree of cottonseed protein hydrolysates was determined by microplate reader, and the result showed that the degree of hydrolysis of CPI hydrolysates was higher than CPG at the same amount of added enzyme, which indicated that CPI was easier to be hydrolysed than CPG. The molecular weight of cottonseed protein hydrolysates was measured by HPLC, and the result showed that the content of peptides (MW≤0.8 kDa) of CPI hydrolysates was significantly higher than CPG with the content of 70%-85% and 40%-60%, respectively. Results of antibacterial activity analysis showed that the antibacterial ability of these protein hydrolysates were strongest when the amount of pepsin-trypsin in digestion simulation system was 5 000-5 000 U (2-2), and the degree of hydrolysis of these protein hydrolysates were (24.72±1.07)% and (19.26±0.39)%, respectively, under such digested conditions, and the ability of both digestions to against E. coli were higher than S. aureus. CPI was higher than CPG in antibacterial activity, while both of the original protein without digestion treatment showed no antibacterial ability. The molecular weight determination results showed that the molecular weight of antibacterial activity peptides of cottonseed protein isolate should be in 0.57-0.75 kDa while cottonseed globulins’ were in 0.66-0.78 kDa.【Conclusion】 There was no significant difference between the two methods in the extracted yield of protein (P＞0.05), proteins classified by Osborne fractionation method showed an obvious difference in subunit composition, but the CPG and CPI showed a similarity. CPI is easier to be digested compared to the CPG and both of these two proteins’ hydrolysates showed an antibacterial activity, the hydrolysates of CPI is higher than CPGs’ in antibacterial activity.

Profile Analysis of the Volatile Flavor Compounds of Quantitative Marinated Chicken During Processing

SUN Zhen, HAN Dong, ZHANG Chun-hui, LI Hai, LI Xia, LIU Zhi-bin, XU Shi-ming

Scientia Agricultura Sinica. 2016, 49(15): 3030-3045. doi:10.3864/j.issn.0578-1752.2016.15.017

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【Objective】This research was conducted to study the compositions and contents of volatile flavor compounds during quantitative marinating process and the taste characteristics of flavor substances in detail. The study was expected to provide theoretical supports to the flavor control technology of quantitative marinating.【Method】 The headspace-solid phase micro- extraction combined with gas chromatography-mass spectrometry (HS-SPME/GC-MS) was used to determine the components and contents of volatile flavor compounds from raw chicken meat, four stages of quantitative marinating process (tumbling, baked 1, steamed, baked 2) and traditional marinating by qualitative and quantitative methods, to calculate odor activity values of the volatile flavor substances by the odor threshold in the water, to determine characteristic flavor substances, main flavor substances and modify flavor substances, then the odor activity value of volatile compounds was analyzed by principal component analysis (PCA) and cluster analysis (CA) procedures.【Result】The results indicated that 54, 60, 60, and 60 kinds of volatile flavor compounds were identified from quantitative marinating process, which were higher than 9 kinds of volatile flavor compounds in the raw and 44 kinds of volatile flavor compounds from traditional marinating. Quantitative analysis revealed that the volatile flavor compounds at tumbling phase mainly from spice liquid. Chicken thigh at baked 1 stage began to appear characteristic flavor substances trans, trans-2,4-decadienal, 1-octene-3-ol and so on, the contents of volatile flavor compounds were the highest at steaming stage, and chicken thigh meat was matured by steaming at this stage, the main stage for the formation of flavor. The contents of total volatile flavor compounds of chicken thigh quantitative marinated at baked 2 stage was much higher than traditional marinating, explaining that quantitative marinating is more intense than traditional marinating in flavor presenting. The characteristic flavor compounds were mainly hexanal, octanal, nonanal, (E)-2-nonenal, (E, E)-2,4-decadienal, (E,E)-2,4-nonadienal, benzeneacetaldehyde, eucalyptol and β-myrcene from quantitative marinated chicken thigh during processing, and the odor activity value of them were significantly higher than that of raw meats and traditional marinated chicken. The main flavor compounds were mainly octanal, (E)-2-decenal, (E, E)-2,4-decadienal, 1-octen-3-ol and β-linalool from quantitative marinated chicken thigh during processing. The modified flavor compounds were mainly ketones, esters and alcohols from quantitative marinated chicken thigh during processing. By principal component analysis and clustering analysis, there were significant differences in odor outline from raw chicken meat, four stages of quantitative marinating process and traditional marinating, and the main flavor compounds clustered into four categories, and each category has characteristic flavor compounds represented quantitative marinating process.【Conclusion】 The flavor profile results showed that compositions and development laws of flavor active substances stewed chicken, comparing with traditional marinating, quantitative marinating can present volatile flavor compounds better.

Lactating Dairy Cows Under Heat Stress Enhanced Gluconeogenesis by Activating the GHIGF-I Axis

LI Lin, AI Yang, XIE Zheng-lu, CAO Yang, ZHANG Yuan-shu

Scientia Agricultura Sinica. 2016, 49(15): 3046-3053. doi:10.3864/j.issn.0578-1752.2016.15.018

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【Objective】Six Holstein cows in early lactation after delivery were chosen to be in heat stress,which were fed 35 days from June 29 in 2013 for detecting milk yield and protein content, growth hormone (GH), insulin like growth factor-I(IGF-I), glucose and liver heat shock protein 70 (HSP70) and gluconeogenesis change in blood to illuminate glucose gluconeogenesis and milk quality decline mechanism when lactating dairy in heat stress from the GHIGF-I axis angle and further revealing the cows heat stress mechanism and control of dairy cows heat stress.【Method】Glucose and GH, and IGF-I contents in blood were detected by collecting jugular venous blood and in vivo taking liver tissue, furthermore milk yield and protein content in lactating cows were counted and analyzed, respectively, before fifth week. In addition, HSP70, PC, PEPCK, GHR, and IGFR were detected by qRT-PCR.【Result】Dairy cows were in heat stress when the average daytime temperature above 32℃ lasted for 25 days and the highest temperature 38℃ lasted for more than 72 h continuously during 35 days feeding period. The milk yield and protein content generally showed a decline with the degree of heat stress in lactating dairy cows deepening from the first week to the fifth week. The expression of HSP70 in the fifth week was significantly higher than that in the first week, so were the expression of PC and PEPCK and GHR and IGFR in liver tissues (P＜0.05) through detecting their contents.【Conclusion】Blood glucose was significantly increased, in the period of study which may be due to the secretion of pituitary GH stimulates the liver to produce more IGF-I, namely through GHIGF-I axis up-regulation of hepatic gluconeogenesis pathway key enzyme expression, gluconeogenesis way was activated with the degree of heat stress in lactating dairy cows deepening. Milk protein content decline may be caused by its precursors was too much for gluconeogenesis to make levels of glucose increase in blood to maintain body’s normal energy supply.

The Detection Methods of the Related Substances in Cefquinome Sulfate Intramammary Infusion

LIU Mao-lin, LIU Yi-ming, XU Fei, ZHANG Dao-kang, HUANG Hui-li, LI Han, LI Xiu-bo

Scientia Agricultura Sinica. 2016, 49(15): 3054-3062. doi:10.3864/j.issn.0578-1752.2016.15.019

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【Objective】 The detection methods of the related substances in cefquinome sulfate intramammary infusion were optimized by referencing the cefquinome sulfate injection in Veterinary Drug Quality Standard Assembly. A specific, sensitive, simple and rapid method was developed, which could be used to analyze the related substance in cefquinome sulfate intramammary infusion, effectively. And it will be useful for its quality control and process optimization. 【Method】 The instrument method was made by the pharmacopoeia requirements. The separation was performed on sodium perchlorate buffer (3.45 g NaClO₄·H₂O was dissolved in 1 000 mL water, mixed in 12 mL phosphoric acid and 115 mL acetonitrile, and adjusted pH to 3.6 with triethylamine) as the mobile phase at flow rate of 1.0 mL·min^-1. The detection wave length was 270 nm and the injection amount of sample solution was 20 µL. The LOD and concentration of the related substance in cefquinome sulfate were estimated by HPLC area normalization method. The linearity, the RSD of injection precision and method precision were tested. For the specificity test, by using 0.2 mol·L^-1 HCL, 0.2 mol·L^-1 NaOH to destroy the drug for 4 h, using 10% H₂O₂ to destroy the drug for 30 min and 90℃ high temperature to destroy the drug for 1 h. The before and after destroyed samples were detected by HPLC after preparation. And the main-substances’ area, related-substances’ concentration, and their resolutions were contrasted with each other, and the number of the peak values of the impurities was also contrasted. 【Result】 After a series of dilutions of the cefquinome sulfate intramammary infusion, when the concerntration of the main component was at 7.5 µg·mL^-1, the S/N of the related-substance was no less than 3, and through calculation of the LOD of the related substance was 0.05 µg·mL^-1, and the RSD of injection precision and method precision of 100 µg·mL^-1 were 0.8% and 0.6%, respectively. The calibration curve of the related substances was a liner equation as y = 45935x-130.03 with a correlation coefficient of 1. The infusions were degraded by almost 10%, 10%, 30%, and 20% under the conditions of acid, alkali, an oxidizing agent, and high temperature, respectively. The retention time and resolution of cefquinome and related-substances were good, and the concentration and numbers of the impurities were added. After detecting the concentration of related substances of the 3 batches of cefquinome sulfate intramammary infusion, and calculated by HPLC method, it was found that the area of related substances were less than 2% of Cef, which met the the Quality Standards of Veterinary Drugs Provisions. 【Conclusion】 The method is simple and easy, selective and sensitive, which can be used to analyze the related substances in agents efficiently.

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