Acta Agriculturae Zhejiangensis

Cloning expression vector construction and expression analysis of tobacco ATPase4 gene

WANG Jing, PENG Shuang, HU Sheng, ZHUO Wei, CHEN Qian, LI Liqin

2019, 31(2): 173-181. DOI: 10.3969/j.issn.1004-1524.2019.02.01

Abstract ( 727 )

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Expression of cytoplasmic membrane ATPase gene in plants can cause cells to release H⁺ to the external environment, thus form the membrane inside and outside electrical chemical formula of the gradient, promote all sorts of ion transport, various stress resistance to the outside world. In order to study the regulatory mechanism of ATPase in tobacco and its related biological functions, this article adopted homologous cloning method to clone a ATPase4 homologous gene from common tobacco K326 and construct ATPase4-pcambia1300 overexpression vector. The bioinformatics software was used to predict the hydrophobic, physical and chemical properties, protein structure, signal peptide, phosphate site and origin of ATPase4. The homology analysis showed that the modified gene had 99% homology with plasma membrane ATPase4 of Nicotiana tomentosiformis, so it was named as plasma membrane ATPase4 gene. Fluorescence quantitative real time PCR (qRT-PCR) was used to determine the relative expression pattern in different tissues under different stresses. The results showed that the length of gene sequence was 2 982 bp, encoding 963 amino acid residues, the gene had the highest expression level in the roots, and it could respond quickly to low potassium, high salt, PEG and cold stress. It was shown that ATPase4 played an important regulatory role in tobacco stress.

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Effects of nitrogen application rate on quality and protein components of low-gluten rice

LAN Yan, HUANG Zhao, HU Mingming, TU Yunbiao, SUN Yingying, SUI Xiaodong, GONG Jing, LI Tian

2019, 31(2): 182-190. DOI: 10.3969/j.issn.1004-1524.2019.02.02

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To investigate the effect of nitrogen application amount on the functional component of low-gluten rice, D105 low-gluten rice was used as material and five nitrogen treatments were set under field cultivation conditions (0, 90, 135, 180, 225 kg·hm^-2), then the effect of different nitrogen levels on the dynamic changes of protein content, processing quality of the grain were investigated and the proportion changes of each component protein were analyzed. The result showed that the total protein content and each component protein content of D105 increased with the increase of nitrogen application amount during grain filling phase. During the maturity period (35 days after flowering), the difference between albumin and gliadin of rice grain was not significant in all fertilization treatments, while globulin and gluten showed different discrepancy, which indicated influence of different nitrogen application amount on albumin and gliadin were slight contrary with globulin and gluten. In addition, different nitrogen application did not change the proportion of component protein. The processing quality and taste quality of D105 low-gluten rice performed properly in the N2 (135 kg·hm^-2)-N3 (180 kg·hm^-2) treatment. When N4 (225 kg·hm^-2) was treated, the integrated quality of rice decreased. In summary, the optimal nitrogen application amount for D105 low-gluten protein was between N2 and N3, and the equation of ratio of gluten to total protein with nitrogen application amount were established. The linear equation: y=6E-05x²-0.02x+39.672 (R²=0.9991). The optimum nitrogen application amount, 166.67 kg·hm^-2, was calculated.

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Characteristics of effective precipitation and water requirement of rice in Guizhou

ZHANG Bo, GU Xiaoping, GU Shuhong, HU Jiamin

2019, 31(2): 191-199. DOI: 10.3969/j.issn.1004-1524.2019.02.03

Abstract ( 643 )

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In order to analyze the characteristics of rice effective precipitation and irrigation water requirement in Guizhou, the daily meteorological data of 80 meteorological stations in Guizhou during 1961-2015 were collected, and the effective precipitation and water requirement in different growth stages of rice in Guizhou were calculated by effective precipitation recommended by the United States Department of Agriculture (USDA) Soil Conservation Agency and reference crop evapotranspiration recommended by the United Nations Food and Agriculture Organization (FAO). The variation characteristics of irrigation amount and irrigation demand index at different rice growth stages were also analyzed. It was shown that the effective precipitation in the whole rice growth season in Guizhou was 324 mm, and presented a decreasing trend. The transplanting-heading period showed the highest effective precipitation along with an increasing trend, yet the effective precipitation in other growth periods showed a decline trend. The total water requirement of the whole rice growth season was 737 mm, and showed a decline trend. Specifically, the water requirement in rice transplanting-heading and heading-filling stage showed a decline trend, yet the water requirement in other growth period showed an increasing trend. The average water shortage during the whole rice growth season was 413 mm along with an increasing trend. The water shortage in rice transplanting-heading stage exhibited a decline trend, yet the water shortage in other growth period showed an increasing trend. The water budget index of the whole rice growht season was 0.56. Within the whole growth season, the rice heading-filling stage was most dependent on irrigation.

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Polymorphism of ELOVL5 gene in Jingyuan chicken

ZHANG Juan, MU Tong, ZHAO Ping, CHEN Jiaping, FENG Xiaofang, GUO Peng, WU Zewen, LIU Liyuan, JIANG Qiufei, GU Yaling

2019, 31(2): 200-206. DOI: 10.3969/j.issn.1004-1524.2019.02.04

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Nucleotide polymorphism and site genetic characteristics of ELOVL5 gene in Jingyuan chicken were studied. Polymorphisms of ELOVL5 gene were detected by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and DNA sequence. The results showed that the ELOVL5 gene exon2 and exon5 had no polymorphism. The total of 1 SNP (g.88620433+1683G>A), 2 genotypes were detected on the intron2 amplified fragment of ELOVL5 gene (AA, AG), genotype AA and allele A was dominant, the frequency was 0.84 and 0.92. No homozygous GG type individual was found in 248 populations of Jingyuan chicken. Population genetic analysis showed that the homozygosity of the locus was higher (0.84) and PIC was 0.14 (PIC<0.25), showing low polymorphism. The results showed that the intron2 mutation of ELOVL5 gene in Jingyuan chicken did not deviate from Hardy-Weinberg equilibrium by χ² test (P>0.05). To sum up, the amplified fragment of the intron2 gene of Jingyuan chicken had low polymorphism, and there was no mutation in exon2 and exon5.

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Isolation and identification of Pseudomonas putida from Leptobotia elongate and pathological lesions of its infection

BAI Minghuan, GENG Yi, DENG Longjun, GAN Weixiong, ZHOU Jian, HUANG Xiaoli, CHEN Defang, OUYANG Ping, ZHAO Ruoxuan, SHEN Bingjie

2019, 31(2): 207-215. DOI: 10.3969/j.issn.1004-1524.2019.02.05

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Pseudomonas putida is a conditional pathogen that widely distributed in water and soil, which can infect aquatic animals such as amphibians, fish and crustaceans. During August in 2017, a serious infection that characterized by ulceration of body surface and bleeding of fins occurred in a loach farm located in Xichang, Sichuan Province of China. To determine the etiology of this outbreak, pathogen isolation, experimental infection, physical and chemical characteristics of isolated bacteria and sequence analysis of 16S rRNA and gyrB gene were carried out. Results showed that a gram-negative, rod-shaped bacillus (designated as BMH170820) was isolated from the diseased fish and its pathogenicity were confirmed by experimental infection test for its pathological changes were consistent with that of natural infection. Based on the colony morphology, physiological and biochemical characteristics, combined with 16S rRNA and gyrB gene sequence analysis, Pseudomonas putida was confirmed as the causative factor of this outbreak. Histopathological observation revealed that Pseudomonas putida infection in Leptobotia elongate were followed as obvious degeneration, necrosis and inflammation in gill, kidney, liver, heart, spleen and digestive tract, the extensive and serious multiple organ dysfunction might lead to the death of Leptobotia elongate. Drug sensitivity tested results showed that this isolated strain of Pseudomonas putida was sensitive to neomycin, ofloxacin, doxycycline, etc, but resistant to amoxicillin, cefalexin and florfenicol.

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Isolation,identification of horse Bacillus cereus and its virulence genes detection

CUI Yilong, SHI Yun, YANG Dahan, YIN Youqin, XUE Jiangdong, HUO Xiaowei, MA Dehui

2019, 31(2): 216-221. DOI: 10.3969/j.issn.1004-1524.2019.02.06

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In order to identify a pathogenic bacterium isolated from the incidence and death racing of an equestrian club in Tongliao city,Inner Mongolia,the isolated strains were subjected to Gram stain microscopy,animal pathogenicity tests,automatic microbiological analysis system detection,16S rRNA gene amplification,phylogenetic tree analysis and partial virulence gene PCR detection. The results showed that the biochemical characteristics of isolates were consistent with the characteristics of Bacillus cereus(B. cereus); it was sensitive to clindamycin, gentamicin and other drugs, moderately sensitive to cefoxitin, ciprofloxacin and other drugs,and resistant to aztreonam,cefazolin and other drugs; 16S rRNA gene sequence was 100% homologous to the registered B. cereus sequence in GenBank; phylogenetic tree analysis showed that the isolated strains were closest to the relatives of B. cereus;the detection results of virulence gene PCR showed that 8 potential virulence genes entFM,cytK,hblA,hblC,hblD,nheA,nheB and nheC were all positive.The results showed that the isolated strain was a serious pathogenic B. cereus.

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Gene cloning and bioactivity analysis of chitinase gene AO-483 from Arthrobotrys oligospora XJ-A1

GONG Shasha, MENG Qingling, QIAO Jun, ZHONG Wenqiang, HUANG Yunfu, ZHANG Guowu, CHEN Ying, CAI Xuepeng

2019, 31(2): 222-228. DOI: 10.3969/j.issn.1004-1524.2019.02.07

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In order to study the functions of chitinase, chitinase gene AO-483 of nematode-trapping fungi Arthrobotrys oligospora XJ-A1 was cloned, analyzed and expressed in prokaryotic expression system Escherichia coli. The pure prokaryotic expression recombinant fusion protein was purified by Ni column and the chitinase activity was determined by DNS method, and the biological activity of recombinant protein AO-483 was verified by dealing with larvae of Caenorhabditis elegans. The results showed that chitinase AO-483 gene encoded 309 amino acids, the homology of its gene sequence was 96.88% and the amino acid sequence was 97.73% respectively with Arthrobotrys oligospora standard strain (ATCC 24927). The chitinase AO-483 belonged to the family 18 glycoside hydrolase with a Glyco-hydro-18 structure domained at amino acid sequence of 20~297 and had two prevalent conserved catalytic domained with the sequence of GMLGG and LDGLDLDVE. The tertiary structure was a typical triosephosphate isomerase barrel structure. The protein identified by SDS-PAGE showed that the recombinant protein had a molecular weight of about 52 ku which was consistent with prediction. Western blot revealed that recombinant protein can specifically react with anti-Arthrobotrys oligospora polyclonal antibody. The activity of recombinant chitinase AO-483 was determined by the DNS method, with a value of 223.31 U·mL^-1. The recombinant AO-483 owned stronger biological activity that can degrade Caenorhabditis elegans larvae stage 1and stage 4.

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Effect of multi-strain co-fermented feed on anti-Salmonella infection in broilers

LIU Jiangying, ZHU Jianjin, JIANG Rong, LIU Ziwei

2019, 31(2): 229-234. DOI: 10.3969/j.issn.1004-1524.2019.02.08

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To investigate the effect of multi-strain co-fermented feed on resisting Salmonella infection in broilers, 50 one-day-old broilers were randomly divided into 5 groups and were fed with different feeds: control group (basal diet+10% unfermented feed), model control group (basal diet+10% unfermented feed), antibiotic control group (basal diet+10% unfermented feed+20 mg·kg^-1 colistin sulfate), 5% fermented group (basal diet+5% unfermented feed+5% fermented feed), 10% fermented feed (basal diet+10% fermented feed). Oral administration of 400 μL Salmonella(1 × 10⁹ CFU·mL^-1) for two days except the control group when the broilers were 13 days old, the control group was given equal volume of sterile water. The broilers were slaughtered at the age of 15 days. Serum levels of pro-inflammatory cytokines (IL-1β, IL-6, TNF-α), endotoxins (LPS) and D-lactic acid (D-LA) were measured. Got colon for HE staining was to observe the shape of colon and got cecum mucosa was to determine the mRNA expression of TLR4 pathway key signaling molecules. The results showed that salmonella infection resulted in a significantly increase level of serum pro-inflammatory cytokines (IL-1β, IL-6 and TNF-α).The mRNA expression of TLR4 signal pathway key signaling molecules (TLR4, MyD88, TRAF6, NF-κB) increased significantly. The level of serum LPS and D-LA increased significantly. While the indexes of antibiotic control group, 5% fermented group, and 10% fermented group were improved to some extent, indicating that the multi-strain co-fermented feed can ameliorate the intestinal inflammation and intestinal barrier destruction that caused by Salmonella infection. The effect was close to 20 mg·kg^-1 colistin sulfate.

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Isolation, culture and identification of adipose-derived stem cells from calves

WANG Shuaishuai, ZHANG Cai, MENG Sudan, HUANG Yongzhi, ZHU Xuemin, WANG Yuqin, YANG Zijun

2019, 31(2): 235-241. DOI: 10.3969/j.issn.1004-1524.2019.02.09

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In order to provide seed cells for related research on vegetative metabolic diseases of cows, adipose-derived stem cells (ADSCs) were eparated from perirenal adipose tissue of calves. ADSCs were collected by organization minced and 0.25% collagenase Ⅰ digestion by different times. The separated cells were proved by the following aspects, observed the morphology, passage 5 of ADSCs were used to detect the surface-specific proteins, passage 5 and 10 were used to detect the growth curve by CCK-8, passage 10 were used to detect the ability of adipogenic and osteogenic differentiation. The results showed that the ADSCs from calves were grown as fibroblast-like after 4 days of inoculation. Immunofluorescence staining showed that the isolated cells expressed Vimentin, CD34, CD44, and CD90, but did not express CD31 and CD45. The cells can be differentiated into adipocytes and osteoblasts under appropriate induction conditions. The separated cells were ADSCs with the ability of adipogenic and osteogenic differentiation.

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β-Conglycinin triggered IPEC-J2 cell damage through p38/JNK MAPK signaling pathway

PENG Chenglu, ZHANG Yu, XIA Xiaodong, HE Mengchu, SHU Yingshuang, FENG Shibin, LI Yu, WANG Xichun, LI Jinchun, WU Jinjie

2019, 31(2): 242-249. DOI: 10.3969/j.issn.1004-1524.2019.02.10

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In this study, in vitro culture techniques were used to study the effects of different concentrations of 7S (β-conglycinin) on IPEC-J2 (intestinal epithelial cells in piglets). The experiment was divided into six groups, A (control), B (1 mg·mL^-1 7S), C (5 mg·mL^-1 7S), D (10 mg·mL^-1 7S), E (5 mg·mL^-1 7S+1μmol·L^-1 SP600125), F (5 mg·mL^-1 7S+1 μmol·L^-1 SB202190). After 24 hours, the viability of the cells was measured by CCK-8 assay. The contents of NO, 5-HT, IL-6 and IL-10 were detected by ELISA, the expression levels of p-JNK, p-p38 and Bcl-2 proteins were detected by western blot, the relative expression of Bad, Bax, Bcl-2 and Caspase-3 mRNA was detected by PCR. The results showed that the viability of IPEC-J2 cells was decreased in groups C and D (P<0.01). The viability of cells in groups E and F was significantly higher than that in group C (P<0.05). 7S promoted the secretion of inflammatory cytokines NO, 5-HT, and IL-6 and decreased the secretion of IL-10, the addition of inhibitors decreased the secretion of cytokines NO, 5-HT, and IL-6, and increased the secretion of IL-10. 7S promoted the expression of p-JNK and p-p38 proteins and decreased the expression of Bcl-2 protein. The addition of inhibitors inhibited the expression of p-JNK and p-p38 proteins and increased the expression of Bcl-2 protein. The ratio of Bcl-2/Bax mRNA decreased with the increase of 7S concentration, and the ratio of Bax/Bcl-x_l increased with the increase of 7S concentration. The relative expression of Caspase-3 mRNA increased with the increase of 7S concentration. On contrary, often adding inhibitors, the ratio of Bcl-2/Bax increased, the ratio of Bax/Bcl-x_l decreased, and the relative expression of Caspase-3 decreased. Our results indicated that 7S caused damage to the intestinal epithelial cells of piglets through the p38/JNK MAPK signaling pathway.

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Rules of sap flow in grapevine during bleeding period

ZHENG Ting, ZHANG Kekun, ZHU Xudong, GUAN Le, SHANGGUAN Lingfei, JIA Haifeng, FANG Jinggui

2019, 31(2): 250-259. DOI: 10.3969/j.issn.1004-1524.2019.02.11

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To investigate the rules of sap flow in grapevine, its inner-and intra-diurnal changes with temperature, humidity, branch length were studied and their correlations were simultaneously analyzed during the bleeding period. In addition, in order to reveal the effects of sap flow on vine growth, relationship among the flows in different vine parts were studied. The results showed that the total bleeding volume ranged from 380 to 540 mL in Rosario Bianco, and the flow lasted for 35-47 d. Flow speed increased first followed by a decrease, and the peak period maintained around 10 d. Branch length greatly affected the bleeding volumes. The sap flow was stable in the longer branch, which mainly contributed to flow; whereas the shorter branch stopped flowing at the earliest time. The flow volume of bleeding sap was stable and evenly distributed in each period. The total bleeding volume varied with bleeding period and differently distributed inner day. During early bleeding period, proportion was gradually increased in the morning and the bleeding volume was larger in the daytime than in the evening. However, the situation was the opposite in the late bleeding period. In addition, we also observed that branch length was directly correlated with flow speed. The flow speed was faster in the longer branch when compared with the shorter ones. The branch position had no significant effect on flow speed. The total flow volume was always greater in branches than trunk. To conclude, the assumed sap flow model was proposed based on the above results, which would lay foundation for the further studies on bleeding sap.

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Effects of lunar phase and temperature on population dynamics of adult Pectinophora gossypiella Saunders

YU Chun, XU Dong, HUANG Wei, LEI Minghong, WAN Peng

2019, 31(2): 260-267. DOI: 10.3969/j.issn.1004-1524.2019.02.12

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Activities of adult pink bollworm were monitored by sex pheromone trap during 2012-2015 in Anxiang County of Hunan Province. A robust linear model based on nonconcave penalized regression was applied to investigate the relationship between the number of trapped moths of pink bollworm and two factors, temperature and lunar phase, and to detect the outliers simultaneously due to the presence of outliers in the collected data. The results indicated that there was a significant linear relationship between lunar phase and activities of adult pink bollworm, while no direct association between lunar gravitation and amount of trapped moths was found from the analysis. The weaker the light at night, the more the number of moths trapped by sex pheromone. Furthermore, the analyzed results illustrated that temperature was a definite inhibitory factor on the trapped moths of pink bollworm. Ranged within 15-35 ℃, as the temperature rises, amount of trapped pink bollworm moths tends to drop markedly.

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Determination of related substances and content of oxyclozanide suspension by high performance liquid chromatography

BAI Yubin, ZHANG Jili, ZHU Jianmin, SU Wenjie, DONG Zhen, ZHOU Xuzheng, ZHANG Jiyu

2019, 31(2): 268-278. DOI: 10.3969/j.issn.1004-1524.2019.02.13

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In order to detect the content and related substances of oxyclozanide suspension, a new method using high performance liquid chromatography (HPLC) was established. Content determination was carried out using Hypersil BDS C18 (150 mm×4.6 mm, 5 μm) column and 0.1% phosphoric acid water-method (volume ratio=38:62) as mobile phase, column temperature was 25 ℃, and determination wavelength was 300 nm. Related substances was carried out using InertSustainSwift C18 (150 mm×4.0 mm,5μm) column, mobile phase, detection wavelength and column temperature were the same as the content determination method. The results showed that liner range of content determination of oxyclozanide suspension were 34.01-104.35 μg·mL^-1 (R²=1.000), and average recovery was 100.1%. The liner range of 2-amino-4, 6-dichlorophenol hydrochloride and 3, 5, 6-trichlorosalicylic acid were 0.51-20.4 μg·mL^-1 (R²=0.996, 1.000, respectively), and average recovery were 110.3% and 98.6%, respectively. It showed that the method was convenient, simple and accurate, and could simultaneously determine the content and related substances of oxyclozanide suspension, and provided standards for quality control of the product.

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Characteristics and formation causes of white horizons in southern China

ZHANG Mingkui, MA Wanzhu, YAO Yucai

2019, 31(2): 279-290. DOI: 10.3969/j.issn.1004-1524.2019.02.14

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To understand the geographical distribution of soils with “albic horizon”, identify accurately the “albic horizon”in the fields and provide the scientific basis for soil investigation and mapping, twenty-five soil profiles with white color horizon were collected from Zhejiang, Fujian, and Guizhou provinces in the southern China. The characteristics of the white color horizon in the region and their relationships with landform and parent materials were analyzed. The results showed that great variations in texture, free iron oxide and organic matter were found in the soils with white color horizon in the southern China. According to the soil color and vertical distribution characteristics of soil free iron oxide, the twenty-five soil profiles were divided into 3 types, i.e., white color horizons affected by parent materials, affected by composite soil processes, and formed by iron removal. The white color horizons affected by the parent materials were not the result of the iron removal, but were related to the low free iron oxide in their parent materials or superimposed parent materials of soil profile. Color for the white color horizons affected by the composite soil processes were not consistent with their free iron oxide profile distribution, as the white color was the result of iron removal in the former soil formation stage, and the iron oxide deposition on the soil structure in the later formation stage covered the iron removal characteristics of the bleach horizon. The white color horizon formed by the action of iron removal was an “albic horizon”, which could be really used for soil classification. The free iron oxide content in the horizon was obviously lower than that of the overlying or the underlying soil. The distribution of the soils with “albic horizon” was most concentrated in the valley plain, gentle slope of hilly mountain and hill ridge, and its micro topography was characterized by the slope and low concave surface, and it was easy to accept the water flow from the height. But their surface generally did not accumulate surface water, with the characteristics of the inner side to the drainage. In order to distinguish from the white color horizon affected by the parent materials, it was suggested that additional diagnostic requirements for the “albic horizon” should be added. The proposed additional diagnostic requirement was that the free iron oxide content of the “albic horizon” was less than 75% of that in its overlying or underlying soil.

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Land use changes induced soil organic carbon stock variations in cropland soils in Fuyang District, Hangzhou City in 1979-2020

QIU Lefeng, YU Zhoulu

2019, 31(2): 291-296. DOI: 10.3969/j.issn.1004-1524.2019.02.15

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Land use change (LUC) is the most dynamic force in terrestrial carbon stock change, and it is imperative to account for the dynamics of LUC in carbon stock change when forming land use policies. In the present study, the impacts of LUCs on carbon (C) stocks were explored at a county scale and the changes of soil C stocks were detected within a county-scale land use planning policy. The LUCs within 1979-2006 in Fuyang and Fuyang Land Use Master Planning (FLUMP) (2006-2020) were selected for this study. The estimates of C stock changes were examined by compiling vegetation and soil organic C density data from six land use types, and through literature reviews and field surveys. The results showed that LUCs between 1979 and 2006 already caused a vegetation carbon (VC) decrease of 273.4 Gg and a soil (0-20 cm layer) organic carbon (SOC) decrease of 771.0 Gg, mainly due to the urbanization processes. Furthermore, the FLUMP (2006-2020) was expected to cause a potential annual C decline of 25.93 kg·hm^-2 for VC and 27.48 kg·hm^-2 for SOC during 2006-2020. As the situation stands, it is urgent to devise rational policies and effective measures to reverse the C loss process.

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Characteristics of nitrogen and phosphorus loss by runoff from paddy and vegetable fields under different fertilization modes in Tiaoxi catchment

LIU Chen, ZHANG Li, LIN Yicheng, GUO Bin, FU Qinglin, LI Hua, DING Nengfei

2019, 31(2): 297-306. DOI: 10.3969/j.issn.1004-1524.2019.02.16

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A continuous two-year field experiment with building runoff ponds on the typical paddy and vegetable lands in Tiaoxi basin was conducted to study the characteristics of nitrogen and phosphorus loss with surface runoff. The nitrogen loss was different between years due to the precipitation. The total nitrogen losses in 2016 and 2017 were 12.76-16.97, 15.40-17.60 kg·hm^-2, respectively, in paddy field, and were 23.06-25.72, 18.11-27.93 kg·hm^-2, respectively, in vegetable field. The total nitrogen loss was mainly in N$O^-₃$-N state in both paddy field and vegetable field. The total phosphorus losses in 2016 and 2017 were 2.36-2.62, 3.57-4.05 kg·hm^-2, respectively, in paddy field. Compared with the conventional fertilization, nitrogen reduction or organic fertilizer did not significantly influence the phosphorus loss. The total phosphorus loss was mainly in soluble phosphorus state in paddy field, whereas it was the particle phosphorus state in vegetable field. The total nitrogen losses were higher in vegetable fields than that in paddy fields, but the total phosphorus losses were the opposite. The fertilizer reduction or organic fertilizer treatments decreased the N loss loading by 7%-35% as compared to the conventional treatment, yet maintained the yield.

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Molecular identification and genetic diversity of a traditional Chinese medicinal fungus Sanghuang

WANG Weike, SONG Jiling, WU Youliang, YAN Jing, LU Na, YUAN Weidong, CHEN Guanping

2019, 31(2): 307-314. DOI: 10.3969/j.issn.1004-1524.2019.02.17

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To study the genetic diversity and genetic relationship of Phellinus baumii, and their relative species by ITS molecular marker technique, and to provide the reference for Phellinus baumii identification and breeding, the ribosomal DNA internal transcribed spacer (rDNA ITS) region was amplified and sequenced to identify the 22 species of Sanghuang which were collected from domestic and detect its genetic diversity. Based on rDNA ITS sequences, the genetic distance and phylogenetic tree were obtained. Results showed that there were distinct genetic divergences among 22 species of Sanghuang, which formed three separate species cluster in the topology tree obviously with high bootstrap value. Meanwhile, two counterfeit Sanghuang strain, Fuscoporia gilva and Inonotus baumii, were detected successfully, and the certified Sanghuang germplasms were distinguished as Sanghuangporus sanghuang unambiguously by BLAST analysis. The results of ITS analysis revealed that Phellinus baumii had the plentiful genetic diversity and the genetic relationships were consistent with morphological characters of taxonomy. ITS method was efficient for identification of Phellinus baumii, which could provide a scientific basis for the resource collection and identification of the species in Phellinus baumii.

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A classification method for hyperspectral imaging of Fusarium head blight disease symptom based on deep convolutional neural network

JIN Xiu, LU Jie, FU Yunzhi, WANG Shuai, XU Gaojian, LI Shaowen

2019, 31(2): 315-325. DOI: 10.3969/j.issn.1004-1524.2019.02.18

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In order to realize rapid and early diagnosis of wheat Fusarium head blight disease via hyperspectral imaging, the correlation between the convolutional layer and the spectrum feature of disease symptom was analyzed, and the classification modeling of hyperspectral image was studied. Two typical neural network, Visual Geometry Group (VGG) and residual neural network (ResNet), were introduced to construct the convolutional neural network with different depth. By comparing the training and testing results of the hyperspectral data set for wheat Fusarium head blight disease, it was shown that the loss value decreased with the increased depth of VGG structure, yet the loss value of validation set was not significantly decreased with the increased ResNet depth. According to the evaluation results for testing set, the VGG network of 4 basic units with 22 layers showed the best performance, as its accuracy of training, validation and testing was 0.846, 0843 and 0.742, respectively. Therefore, the VGG network could effectively extract the spectrum feature of Fusarium head blight disease. These results would provide theoretical basis for the intellectual diagnosis of wheat Fusarium head blight disease by the remote sensing in a large scale.

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Wheat germ sheath recognition based on image features

YAN Jianwei, SU Xiaodong, ZHAO Yuan, LIU Jinping

2019, 31(2): 326-332. DOI: 10.3969/j.issn.1004-1524.2019.02.19

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Using color features of the image, wheat seed image was firstly divided and wheat seed contour moment was determined. The centroid coordinates of wheat seed were determined according to the contour distance. Then the coleoptile image was divided according to color image of wheat coleoptile image to obtain wheat coleoptile. Followed by Zhang-Suen parallel refinement algorithm, wheat coleoptile was refined to obtain the coleoptile skeletal line, and then obtain all the points on the skeletal line image (single pixel) to perform a multi-segment linear curve approximation of the coleoptile skeletal line. Finally, orientation of the coleoptile and position of the cut point of wheat coleoptile were determined based on centroid coordinates of wheat seed outline, approximate curve of the coleoptile skeletal line, and cutting distance (given). Image processing of 30 pairs of wheat coleoptiles was verified by image processing. The results showed that this method could accurately segment wheat coleoptile image, quickly identify wheat coleoptile, attitude and other information. Identification and localization of wheat coleoptile based on image color features provided an accurate, rapid and visual method for the identification, extraction and analysis of wheat coleoptile, and it had great significance for the construction of a visual system and automatic recognition of germ sheath.

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Research progress on effects of straw returning on nitrogen cycling microbes and functional genes in paddy soil

WANG Qingxia, CHEN Xijing, YU Man, SHEN Alin

2019, 31(2): 333-342. DOI: 10.3969/j.issn.1004-1524.2019.02.20

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The response of soil microorganisms to soil changes is extremely sensitive, and the impact of straw returning on soil microbial community is important for the soil nitrogen cycling maintainance in paddy field systems. Straw incorporation to soils influences the biochemical transformation of soil nitrogen by changing the soil microecological environment. The nitrogen management strategy in rice production is closely related to these changes in soil nitrogen transformation. Based on the latest published research papers, this study reviewed the soil nitrogen transformation processes, the functions of related functional microbes, and the driving mechanisms of the key functional genes in paddy field soil with straw incorporation. It would help to understand the microbial mechanism of soil nitrogen cycling in paddy field soils with straw incorporation, and provide theoretical basis for the scientific application of nitrogen fertilizer in paddy field systems.

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