Journal of Henan Agricultural Sciences

Contents

2020, 49(6): 0.

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Research Progress of the Formation Mechanism of Apomictic Seed in Chinese Chive

ZHANG Huamin, CHEN Jianhua, YIN Shouheng, MA Aichu, ZHANG Ming, XIAO Wanlu, JIAO Ziyuan

2020, 49(6): 1-7. DOI: 10.15933/j.cnki.1004-3268.2020.06.001

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In this paper,the formation mechanism of apomictic seed in Chinese chive was reviewed from the meiosis of megaspore mother cell and microspore mother cell,the development of embryo sac and pollen grain,the embryonic development and seed formation,and the heredity of apomixis. In the mean time,the research direction of apomictic characteristics and its application prospect in Chinese chive breeding were prospected,with a view to provide reference for the clarification of apomixis mechanism and the study of genetics and breeding of Chinese chive in the future.

Transcriptome Analysis of Wheat Roots in Response to Heavy Metal Pb Stress

WANG Yiren, NIE Mengjie, WANG Yuquan, HU Xigui, DING Weihua, HU Tiezhu

2020, 49(6): 8-15. DOI: 10.15933/j.cnki.1004-3268.2020.06.002

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In order to explore the molecular mechanism of wheat in response to heavy metal Pb stress,the wheat cultivar AK58 was treated with different concentrations [0 (CK),40,80,160 mg/L] of Pb by hydroponic method, the root transcriptome was sequenced, then the differentially expressed genes were screened and analyzed, and GO classification and KEGG enrichment analysis were done.The results showed that under different concentrations of Pb,the root length and number of wheat were inhibited,and the inhibition effect was stronger with the increase of Pb concentration.A total of 38 904 differentially expressed genes were obtained between Pb stress treatment and CK.Among them,there were 6 072,16 581 and 16 251 differentially expressed genes under 40,80 and 160 mg/L Pb stresses,respectively.The differentially expressed genes of treatment with 80 mg/L Pb were selected as the research focus for GO classification and KEGG pathway enrichment analysis. GO classification showed that up-regulated differentially expressed genes were mainly involved in immune system processes,exercise processes,metabolic processes,rhythmic processes,catalytic activity and electron carrier activity;the down-regulated differentially expressed genes were mainly involved in development,growth,localization,replication,reproduction processes,transport activity and antioxidant enzyme activity.KEGG enrichment analysis found that up-regulated differentially expressed genes were mainly involved in plant hormone signal transduction pathway,plantpathogen interaction pathway,drug metabolism pathway and MAPK signaling pathway;the down-regulated differentially expressed genes were mainly involved in the secondary metabolite biosynthesis pathway,phenylpropane biosynthesis pathway,antibiotic biosynthesis pathway,carbon metabolism and sucrose metabolism.Six differentially expressed genes responding to Pb stress were selected for RT-PCR verification.The results showed that the expression patterns of the six differentially expressed genes were consistent with the results of RNA-Seq analysis,which further verified the accuracy of the RNA-Seq results.

Effects of Reduced Application Rate of Nitrogen on Yield and Economical Benefit of Wheat under Different Tillage Methods

NIE Shengwei, ZHANG Haoguang, ZHANG Qiaoping, XU Jidong, ZHANG Yuting

2020, 49(6): 16-22. DOI: 10.15933/j.cnki.1004-3268.2020.06.003

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The economical benefits of wheat of EF(CK,nitrogen application rate of 300 kg/ha),RF10(reducing 10% of nitrogen fertilizer),RF20( reducing 20% of nitrogen fertilizer) and RF30( reducing 30% of nitrogen fertilizer)treatments were studied under vertical rotary tillage [FL,(30±5) cm] and traditional rotary tillage [XG,(12±5) cm] methods in 2017—2018 and 2018—2019 respectively,so as to provide reference for reducing nitrogen fertilizer application and efficiently utilizing resources.The results showed that,compared with FL-EF treatment,FL-RF20 treatment did not decrease the wheat yields in two wheat seasons;FL-RF20 treatment obtained higher net income and ratio of output to input,while FLRF10,FL-RF30 treatments decreased net income and ratio of output to input in the first wheat season,and increased in the second wheat season,respectively. The net incomes of FL-EF,FL-RF10,FL-RF20,FLRF30 treatments were 8 030. 5,7 342.4,8 891.7,5 742.9 Yuan/ha in 2018,and 13 791.5,14 231.2,14 616.8,13 823.8 Yuan/ ha in 2019,respectively.Compared with XG-EF treatment,XG-RF20 treatment did not decrease the wheat yields in two wheat seasons;XG-RF20 treatment obtained higher net income and ratio of output to input,while XG-RF10,XG-RF30 treatments decreased net incomes in two wheat seasons.The net incomes of XG-EF,XG-RF10,XG-RF20,XG-RF30 treatments were 9 190.0,8 832. 5,9 714.7,7 205.4 Yuan/ha in 2018,and 13 452.6,13 257.1,15 022.4,12 465.6 Yuan/ha in 2019,respectively.Compared with traditional rotary tillage method,total input of vertical rotary tillage method increased by 375.0 Yuan/ha,but net income of vertical rotary tillage method increased by 217.7 Yuan/ha.Overall,vertical rotary tillage method is better than traditional rotary tillage method,which could increase the soil layer depth,improve soil structure,adjust soil function,and promote sustainable productivity.

Effects of Growth Environment and Culture Medium on Haploid Embryo Germination Rate and Plantlet Regeneration Rate of Wheat

WU Xiaojun, HU Xigui, CHEN Xiangdong, JIANG Xiaoling, HU Tiezhu, LI Xiaohui, CHENG Wei, RU Zhengang

2020, 49(6): 23-29. DOI: 10.15933/j.cnki.1004-3268.2020.06.004

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In order to improve the germination rate and plantlet regeneration rate of wheat haploid embryos,the induction rate and plantlet regeneration rate of haploid embryo from hybrid spike of different wheat materials × maize under different growth environments (culture in vitro, pot culture and field culture)and different media[1/2MS medium,named MC1; adding 50 mg/L vitamin C and 150 mg/L glutamine in 1/2MS medium,named MC2;adding 0.5 mg/L IAA (indole-3-acetic acid),0.5 mg/L KT(kinetin),150 mg/L glutamine in 1/2MS medium,named MC3]were analyzed.The results showed that environmental condition and culture medium had significant effects on the induction rate and plantlet regeneration rate of haploid embryo. The quality of haploid embryo obtained from wheat hybrid spike cultured in vitro was higher,the germination rate and plantlet regeneration rate of haploid embryo were 81.5% and 63.0%,respectively,which were 60.1% and 3.4 times higher than those from field culture,30.4% and 68.0% higher than those from pot culture.The culture effects of different wheat materials were the best on MC3,the average germination rate and plantlet regeneration rate of wheat haploid embryo were 42.3% and 15.3% on MC1,46.2% and 27.5% on MC2,and 65.6% and 38.2% on MC3,respectively.In summary,1/2 MS+0.5 mg/L IAA+0.5 mg/L KT+150 mg/L glutamine had good culture effect on the small and medium-sized embryos and immature embryos with poor development,and obtained relatively high germination rate and plantlet regeneration rate,but the rate of one-time seedling formation was low,so it was necessary to replace the medium for root-promoting seedling culture.

Effects of Modified Diammonium Phosphate on Growth,Photosynthetic Physiological Characteristics and Yield of Mung Bean

LIU Ya, CAI Guangrong, MU Baomin, LI Bing, WANG Shiya, LIU Meiling, ZHENG Dianfeng, FENG Naijie

2020, 49(6): 30-41. DOI: 10.15933/j.cnki.1004-3268.2020.06.005

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In order to provide a theoretical basis for the application of modified phosphate fertilizer double active diammonium which mainly included diammonium phosphate and lignin in mung bean production,four phosphorus levels,namely P0(no phosphorus),P1(diammonium phosphate,225 kg/ha),and P2(double active diammonium,225 kg/ha) and P3(1/2 double active diammonium,112.5 kg/ha)were applied on Lüfeng 2 and Lüfeng 5 mung bean genotypes.Mung bean dry weight,root morphology,photosynthetic physiological characteristics and yield were calculated.The results indicated that both P2 and P3 treatments increased the plant dry weight of mung bean during V3—R6 stage compared to the P0 and P1 treatments,and the maximum value appeared in P3 treatment at R6 stage. Compared with the P0 and P1 treatments,Lüfeng 2 aboveground dry weight increased by 57.55% and 34.31%,and the root dry weight increased by 19.63% and 11.84%.The performance of Lüfeng 5 was similar to that of Lüfeng 2,with 30.29% and 21.62% increase in aboveground dry weight,and 36.66% and 34.56% increase in root dry weight.P2 and P3 treatments significantly increased mung bean total root length,total root surface area at R4 stage,increased average root diameter at V5 stage and total root volume at R4 stage,and significantlyenhanced net photosynthetic rate of Lüfeng 2 and Lüfeng 5 at R6 stage. Both P2 and P3 treatments increased plant soluble protein content of Lüfeng 2 at R4 stage and decreased simultaneously it at R6 stage.They also increased leaf soluble protein content of Lüfeng 5 at R4 stage and significantly decreased acid phosphatase activities in the leaf and root at R2 and R6 stages,respectively.P2 and P3 treatments could increase stem thickness of Lüfeng 2 and plant height of Lüfeng 5,and also increase the number of grainsper plant,the hundred grains weight and the weight of grains per plant of two genotypes. The number of grains per plant,the hundred grains weight and the weight of grains per plant of P3 treatment in Lüfeng 2 were 14.85% and 48.28%,15.41% and 13.26%,16.94% and 44.16% higher than those of the P0 and P1 treatments,respectively;and those of P3 treatment in Lüfeng 5 were 46.51% and 11.35%,3.34%and 8.70%,51.20% and 28.20% higher than those of the P0 and P1 treatments,respectively.It can be seen that modified diammonium phosphate can effectively promote the growth of mung bean plants,and improve plant stress resistance by increasing the content of soluble protein,promote the accumulation of photosynthetic products at the later stages by increasing the photosynthetic capacity,and finally increase the mung bean yield.

Effect of Molybdate Fertilizer on Enzyme Activity in Tobacco Growing Soil,Yield and Quality of Flue-cured Tobacco

SHEN Hongtao, DUAN Weidong, LI Dong, WANG Xinzhong, ZENG Qiang, XU Chensheng

2020, 49(6): 42-47. DOI: 10.15933/j.cnki.1004-3268.2020.06.006

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In order to explore the effects of the amount of molybdate fertilizer on soil enzyme activity and yield and quality of flue-cured tobacco,a field experiment was conducted,with flue-cured tobacco variety K326 as experimental material.The treatments were comprised of four ammonium molybdate application levels:0,45,90 and 135 mg/plant ammonium molybdate,which were denoted as CK,T1,T2 and T3,respectively.The effects of ammonium molybdate on soil enzyme activity, conventional chemical composition of tobacco leaves, sensory quality and economic characters of flue-cured tobacco were studied.The results showed that the application of ammonium molybdate significantly increased the activities of urease,sucrase and catalase in soil,and the increase amplitudes were 17.51%—46.79%,9.67%—25.21% and 8.56%—55.93%, respectively.With the increase of ammonium molybdate application amount,the content of reducing sugar,total nitrogen,potassium and the ratio of reducing sugar to total sugar of flue-cured tobacco leaves increased significantly, while the content of nicotine and chlorine of flue-cured tobacco leaves first decreased and then increased.The application of ammonium molybdate could increase aroma quality,aroma quantity,concentration and tender degree of flue-curedtobacco leaves,and increase the yield,output value and proportion of superior and moderate flue-cured tobacco. Compared with CK,T1,T2 and T3 treatments increased the output value of flue-cured tobacco leaves by 4.63%,13.07% and 8.70%,respectively,and the proportion of superior and moderate tobacco increased by 3.07%, 7.39% and 3.95%, respectively.In conclusion, the application of ammonium molybdate could significantly improve the activities of soil enzymes and the soil ecological environment,coordinate the internal chemical composition, increase the sensory quality of tobacco leaves and the economic benefit of flue-cured tobacco.Under the condition of this experiment,the application amount of 90 mg/plant ammonium molybdate was the best option in Nanping tobacco area.

Effects of Cd Stress on Bacterial Community Composition and Diversity in Potato Rhizosphere Soil

XIE Zhao, FU Tianling, HE Tengbing, CHENG Jianbo, FU Dongshun, LUO Sheng

2020, 49(6): 48-58. DOI: 10.15933/j.cnki.1004-3268.2020.06.007

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In order to reveal the difference of bacterial community structure in rhizosphere soil and the change characteristics of rhizosphere bacteria under Cd stress,and understand the relationship between bacteria and Cd tolerance in potato rhizosphere soil,the bacterial diversity and community structure of potato rhizosphere soil under Cd stress with different concentrations(0,10,50,100 mg/kg)were studied by high throughput sequencing.The results showed that the bacterial diversity of potato rhizosphere soil was the lowest when the Cd stress concentration was 100 mg/kg,and the highest when the Cd stress concentration was 10 mg/kg.Proteobacteria was the dominant phylum at phyla level, accounting for 37.78%—48.28%.At genus level, Mizugakiibacter was dominant, accounting for 9.16%—20.44%.Principal component analysis showed that the community structure of bacteria in potato rhizosphere soil under Cd stress was significantly different from that of control.Three genera of Cd-tolerant bacteria,Brubacter,Flexivirga and Jatrophilhabitans,were identified by correlation analysis.

Comparison of Potassium Solubilization Ability and Drought Resistance of Five Potassium-Solubilizing Bacteria

FENG Xiaoying, ZHOU Guixiong, WU Qingshan, FANG Zheng, WANG Yuqian, NIU Xiaojuan, WENG Qingbei

2020, 49(6): 59-63. DOI: 10.15933/j.cnki.1004-3268.2020.06.008

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In order to screen potassium-solubilizing bacteria suitable for the preparation of biological fertilizers in arid regions,potassium feldspar powder was used as the sole potassium source,the potassiumsolubilizing ability of five bacteria strains (LG1,LG8,WG33,WG36 and WG40) was explored,and an experiment was conducted under simulated drought condition to explore the effects of inoculating fermentation broth on the drought resistance of maize seedlings. Results showed that,all the five strains had potassium-solubilizing ability,sequenced as WG33>LG1>WG40>WG36>LG8,WG33 ranked the first,and the soluble potassium content in the fermentation supernatant reached 7.73 mg/L.Under drought condition, inoculating five potassium-solubilizing bacteria enhanced the plant height of maize seedlings,decreased the MDA content and increased POD,SOD activities, indicating that the five potassium-solubilizing bacteria strains can improve the drought resistance of maize seedlings.

Research on Diversity and Plant Growth Promoting Activities of the Cultivable Rhizosphere and Endophytic Bacteria of Miscanthus

PANG Fahu, XU Ge, LI Min, XIE Hanzhu, CHEN Zhaojin

2020, 49(6): 64-73. DOI: 10.15933/j.cnki.1004-3268.2020.06.009

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In order to understand the diversity of cultivable rhizosphere and endophytic bacteria associated with Miscanthus,strains with different colony morphologies were isolated using the dilution plate counting method and their plant growth-promoting activities were tested. Phylogenetic analysis based on partial 16S rRNA gene sequences indicated that 86 strains could be categorized into 4 phyla (Actinobacteria,Bacteroidetes,Firmicutes and Proteobacteria),7 classes,and 22 genera.The bacterial community compositions in the rhizosphere and plant tissues were different from each other.The dominant genus Curtobacterium was found to be well adapted to the rhizosphere,as well as to the root,stem and leaf.The dominant genera in the rhizosphere were Bacillus and Rhizobium,that in the root was Burkholderia,and in the stem was Sphingomonas.Plant growth-promoting activities analyses indicated that most strains displayed plant growth-promoting effects including IAA-production,phosphorus-solubilization,and arginine decarboxylaseproduction,and that the dominant plant growth-promoting bacteria were Curtobacterium,Rhizobium,Bacillus,Burkholderia and Sphingomonas.Plant growth-promoting activities and bacterial community composition were different between rhizosphere and plant endospheres.

Characteristics of Nitrogen Losses of Organic Fertilizers in Purplish Soil in Simulating Runoff Experiments

XIA Hongxia, ZHU Qihong, LIU Xidong, LI Qiang, WANG Shumin, DING Wuquan, YANG Zhimin, CHEN Yucheng

2020, 49(6): 74-83. DOI: 10.15933/j.cnki.1004-3268.2020.06.010

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In order to study the differences of nitrogen loss between organic fertilizers and chemical fertilizerafter application in purplish soil,according to local agricultural habits in Chongqing,chemical fertilizer,swine manure (SM),oil cake (OC),biogas slurry (BS) and organic-inorganic compound fertilizer(OF) were selected to compare the characteristics of nitrogen loss by indoor simulating runoff experiment in purplish soil.The results showed that,under the experimental condition,total nitrogen contents in runoff fluid of fertilizer treatments were all significantly(P<0.01) higher than blank,with 3.4—9.1 times,while the differences between organic fertilizers and chemical fertilizer were insignificant(P>0.05).The order of total nitrogen loss rates of organic fertilizers relative to chemical fertilizer was SM>OF>BS>OC.The rainfall intensity,fertilizing amount and slope had significantly positive relation with runoff total nitro gen(TN) content in purplish soil (P<0.05),and slope was the most important factor for causing nitrogenloss,followed by fertilizing amount and rainfall intensity.Under the indoor simulating condition,particulate nitrogen was the main form in runoff, accounting for 52.51%—81.50% of TN, ammonium nitrogen(NH₄⁺-N) was higher than nitrate nitrogen (NO₃^--N),NH₄⁺-N occupied 11.44%—43.57% of TN and NO₃^--N only occupied 1.74%—14.55%. It presented that organic fertilizer application would cause nitrogen loss by runoff water in purplish soil,and nitrogen loss of organic fertilizer might be significantly higher than that of chemical fertilizer in a certain condition.Thus,different kinds of organic fertilizers should be applied reasonably in agricultural production,and fertilizing amount should be controlled.

Identification of Adult-Plant Resistance to Stripe Rust and Molecular Marker Detection of Yr Gene in 242 Wheat Varieties(Lines)

ZHOU Jun, LI Kuiyin, ZHANG Li, PENG Qin, XU Ruhong, REN Mingjian

2020, 49(6): 84-97. DOI: 10.15933/j.cnki.1004-3268.2020.06.011

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Two hundred and forty-two wheat germplasms from Guizhou,Sichuan and other places were collected,the adult-plant resistance to stripe rust was identified in the field,and five known resistance genes(Yr5,Yr10,Yr15,Yr18,Yr26) markers were used to detect the materials,so as to determine the stripe rust resistance level and type of disease-resistant genes of wheat germplasms,which would lay the foundation for the scientific and reasonable use of excellent disease-resistant wheat germplasms.The results showed that 116 wheat germplasms at adult stage displayed resistance to stripe rust among 242 wheat germplasms,including 11 germplasms were immune,27 germplasms were nearly immune,41 germplasms were highly resistant and 37 germplasms were moderately resistant to stripe rust,which accounted for 4.54%,11.16%,16.94% and 15.29% of the germplasms,respectively;only one germplasm exhibited slow rust,accounting for 0.41%;125 germplasms exhibited medium sensitivity and high sensitivity,accounting for 51.65%.Molecular detection result showed that,59 germplasms carried Yr26,13 contained Yr18,21 contained Yr15,14 contained Yr10,and 52 contained Yr5.There were 39 germplasms containing 2 resistant genes,such as Guinong No.19;7 germplasms carried 3 resistant genes,such as Guinong 775 and Guixie No.3;80F-1-4-2 contained 4 resistant genes.The above Yr genes were not detected in the remaining disease resistant germplasms,which might carry other undetected disease resistant genes.

Effects of Seed Coating Agents on the Occurrence of Main Pests and Maize Yield in Corn Field

ZHAO Man, TANG Jinrong, DONG Shaoqi, WANG Xinhui, MIAO Changjian, GUO Xianru

2020, 49(6): 98-107. DOI: 10.15933/j.cnki.1004-3268.2020.06.012

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In order to clarify the influence of major neonicotinoid seed coating agents on the occurrence of important pests and plant yield in maize field,and select safe and effective seed coating agents that are suitable for controlling main pests in maize field and increase maize yield,this study evaluated the effects of four flowable concentrate seed coating agents (600 g/L imidacloprid, 30% thiamethoxam, 200 g/L clothianidin and 48% clothianidin·imidacloprid)on maize emergence rate,occurrence of target pests and non-target pests,and maize yield under field conditions.The results indicated that the tested neonicotinoid seed coating agents had no significant effects on maize emergence rate and showed a good control effect on target pests including Laodelphax striatellus,Rhopalosiphum maidis and Aphis gossypii.Furthermore, 48% clothianidin· imidacloprid FSC had the best control effect against target pests among the four tested neonicotinoid seed coating agents and its control efficacies to target pests such as R.maidis and A.gossypii were still more than 90% at the later growing stage of maize.The population density of Pseudaletia separata in 30% thiamethoxam FSC treated plot at maize late filling stage and mature stage was remarkably higher than control(treated by clean water)plot,while the population density of non-target lepidopteran pests in other seed coating areas was equivalent or significantly lower than that in control plot. For the maize yield,the seed coating agents treated plots all increased to a certain extent when compared with the control area,and the 48% clothianidin· imidacloprid FSC treated plot increased largest with an increase rate of 10.03%.All the above results indicated that 48% clothianidin·imidacloprid FSC could be used to treat maize seeds in maize production since it not only had significant control efficacy to piercing-sucking pests through the whole growth period of maize,but also could increase the corn yield.

Analysis of Cytochrome b(Cyt b) Gene Sequence from Low Sensitivity Rhizoctonia solani Isolates to Azoxystrobin

FANG Yali, ZHANG Zhijia, JIA Jingjing, ZHOU Yaoyao, XING Kun, HAN Pengjie

2020, 49(6): 108-114. DOI: 10.15933/j.cnki.1004-3268.2020.06.013

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In order to explore molecular mechanism of R.solani low sensitivity isolates to azoxystrobin and provide reference for integrated control of potato black scurf disease in field,the sensitivity of fifteen Rhizoctonia solani isolates to azoxystrobin and partial isolates to azoxystrobin with SHAM was detected by growth rate method,then the sequence structure and mutant sites of Cyt b related to azoxystrobin resistance were analyzed in seven low sensitivity isolates and one high sensitivity isolate,and the sensitivity of R.solani to four botanical fungicides,osthole,ethylicin,matrine and amino-oligosaccharin was detected by growth rate method.The results indicated that only H15 isolate showed high sensitivity to azoxystrobin in the tested isolates,and EC₅₀ value were 0.49 μg/mL.The rest 14 isolates showed low sensitivity to azoxystrobin,and EC₅₀ values were above or equal to 32 μg/mL.The sensitivity of R. solani isolates to azoxystrobin was increased after SHAM was added,however,EC₅₀ values of low sensitivity isolates H5 and H16 were still above 32 μg/mL.Sequence analysis showed no point mutation at codons 129,137 and 143,and absence of intron immediately downstream of codon 143 in the 8 isolates with different sensitivity to azoxystrobin.The EC₅₀ values of osthole,ethylicin,matrine and amino-oligosaccharin were 4.57,7.72,15.81 and 116.58 μg/mL to R.solani isolate,respectively.The sensitivity of R.solani isolates to azoxystrobin varies largely,and low sensitivity R.solani isolates to azoxystrobin may have alternative oxidation pathway besides alternative respiration pathway inhibited by SHAM in vitro.The absence of intron immediately downstream of codon 143 suggests that R.solani may easily produce resistance to azoxystrobin.The inhibition effect of osthole and ethylicin on R.solani is better,so these two botanical fungicides could be used to control potato black scurf as alternative pesticides.

Effects of Artificial Nutrient Substrate on Acidification and Accumulation of Phenols in Continuous Cropping Cucumber Soil

YU Chaoge, LÜ Tinghui, XIE Zhihui, HE Ruifa, YAO Mengdie

2020, 49(6): 115-119. DOI: 10.15933/j.cnki.1004-3268.2020.06.014

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In order to know the effect of artificial nutrition substrate on continuous cropping obstacle,contents of benzoic acid,p-hydroxybenzoic acid,ferulic acid,vanillin,total phenols and pH were detected in the soil and nutrition substrate with cucumber continuous cropping for 1—19 times.Cucumber seed germination rate was also investigated after treated with water extract of above cultivation medium.Results showed that contents of benzoic acid,p-hydroxybenzoic acid,ferulic acid and total phenols all rose,while pH decreased with the increase of cucumber continuous cropping stubble times.Artificial nutrition substrate alleviated the phenols content rising and pH decreasing.And this effect generally began to produce after continuous cropping for 7 times or 9 times. Meanwhile,nutrition substrate also reduced inhibitory effect of soil water extract on seed germination. That continuous cropping obstacle is alleviated by nutrition substrate may have relation with less phenols accumulation and pH value decrease than soil.

Study on Genetic Diversity of Actinidia Germplasm Resources Based on SCoT Markers

ZHOU Yuqiao, GUO Guoye, ZHOU Suo, DU Ge, LI Shulin

2020, 49(6): 120-126. DOI: 10.15933/j.cnki.1004-3268.2020.06.015

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SCoT molecular markers were applied to 28 Actinidia germplasm resources to study genetic diversity and UPGMA clustering relationship.The results showed that,8 polymorphic SCoT primers were selected,a total of 75 clear bands were obtained from 28 Actinidia materials with 8 primers,and 9.1 polymorphic bands were obtained from each primer on average,with a polymorphic ratio of 97.3%.The genetic similarity coefficient ranged from 0.47 to 0.86,indicating that the tested Actinidia germplasm resources had abundant genetic diversity.UPGMA clustering analysis showed that 28 Actinidia germplasms could be divided into three distinct clustering groups at the genetic similarity coefficient of 0.64,and part of A.chinensis,two A.arguta germplasms and most germplasms of A.chinensis var.deliciosa clustered together,which showed that A.chinensis, A.arguta and A.chinensis var.deliciosa had high genetic similarity and closely genetic relationship among these germplasms.The genetic variation of some A.chinensis germplasms was high and had closely genetic relationship with A.macrosperma.

Effects of Reduced Fertilization on Yield,Nutrient Uptake and Soil Nutrient Residue of Grape

LÜ Zhongwei, GUO Zhanling, ZHANG Ke, WU Wenying, LOU Yusui, YANG Zhanping

2020, 49(6): 127-131. DOI: 10.15933/j.cnki.1004-3268.2020.06.016

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In order to realize the sustainable development of green ecological production of grape,four fertilization modes,CF(farmers’ habitual fertilization,three times application),RF(30% reduction of CF,three times application),SF(nitrogen,phosphorus reduction and potassium increase of CF,two times application),RSF(30% reduction of SF,two times application),were set up by field experiments to study the effects of reducing fertilization amount and frequency on Shine Muscat grape yield,nutrient uptake and soil nutrient residue. The results showed that RF, SF and RSF could increase the fresh fruit yield compared with CF and decrease the water content of grapes, increase the absorption of nitrogen,phosphorus and potassium in leaves,reduce the residue of nitrate nitrogen in one meter soil profile.SF and RSF could significantly increase soil available potassium residue in 60 cm soil profile.Among them,RF was the best fertilization mode,which increased the fresh weight of grape fruit by 3.1%,decreased the moisture content by 1.0 percentage points,increased the absorption of nitrogen,phosphorus and potassium of leaves by 13.9%,22.7%,14.0%,increased the absorption of potassium of fruit by 2.5%,decreased nitrate nitrogen residue in 1-meter soil by 16.4%,and decreased available phosphorus and potassium residues by 16.6% and 12.7% in 60 cm soil profile.Therefore,reasonable reduced fertilization can not only reduce fertilizer input and production cost,but also increase grape yield and nutrient absorption,reduce soil nutrient residue,and can be applied in production.

Effect of Rejuvenating Trunk Buried Combined with IBA Treatment on Apple Cuttings Rooting

HUANG Song, LI Ming, JIAO Jian, SONG Chunhui, WANG Miaomiao, BAI Tuanhui, WANG Hao, FENG Jiancan, ZHENG Xianbo

2020, 49(6): 132-136. DOI: 10.15933/j.cnki.1004-3268.2020.06.017

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In order to solve the problem of low rooting rate of apple cuttings,one year old Fuji,Melba and Malus robusta Rehd cuttings were used as materials to study the effects of rejuvenating trunk buried combined with IBA treatment on rooting.The results showed that the rooting rate,rooting number,root length and rooting index of rejuvenated trunk of Fuji,Melba and Malus robusta Rehd cuttings that grew in the medium with cow dung and loam mixed in a ratio of 2∶3 were all higher than those of the nonrejuvenated trunk. With the increase of IBA concentration,the four indexes all showed the trend of first rising and then declining.When IBA concentration was 1 250 mg/L,the rooting rate,rooting number,root length and rooting index were the highest.The rooting rates were 91.8%, 90.8% and 93.1%,respectively.The rooting numbers were 25.8,45.8 and 39.2.The root lengths were 60.6,60.3,48.8mm.The rooting indexes were 145.0,250.2 and 177.7.This study found that combination of rejuvenating trunk buried with IBA treatment could greatly improve the rooting rate of apple cuttings and achieve rapid and efficient cloning of apple cultivar.

Production and Characterization of PCV2b Epitope Chimeric Virus-like Particles Expressed in E.coli

FENG Hua, LIU Qingkun, REN Chunxiao, LIU Yunchao, ZHANG Gaiping

2020, 49(6): 137-142. DOI: 10.15933/j.cnki.1004-3268.2020.06.018

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To develop a high efficient and economic PCV2b subunit vaccine,the codons of PCV2 Cap gene were optimized for E.coli expression system,and the gene sequence of PCV2 neutralizing epitope ²²⁶LKDPPLNP²³³was conjugated at its C-terminus(Capc).After the recombinant sequence was constructed on PE-Sumo vector,recombinant chimeric Capc was expressed in E.coli.By using SDS-PAGE,Western blot,Dynamic light scattering and TEM,the immune reactivity and the structure of Capc protein were preliminarily identified.The results showed PE-Sumo-Capc vector was successfully constructed and transferred into E.coli BL21(DE3);Sumo-Capc protein was mostly expressed in soluble form after 12 h induction under 26℃ .The final purified Capc protein was able to specifically react with PCV2 Cap monoclonal antibody,and self-assembly into virus-like particles with the diameter around 18 nm.The current study successfully constructed an E.coli expression system for PCV2b neutralizing epitope chimeric Cap protein,which showed a good immunogenic reactivity and could self-assembly into virus-like particles.

Screening of Up Regulated Genes Related to Follicle Development Based on Illumina Sequencing in Cattle

MENG Jinzhu, AN Qingming, WU Yuanxun, ZHAO Yuanyuan

2020, 49(6): 143-149. DOI: 10.15933/j.cnki.1004-3268.2020.06.019

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To study the important regulatory factors promoting follicle growth into dominant follicle and their expression pattern in bovine follicular development,the largest follicle(8—10 mm) and the second largest follicle(5—8 mm)were acquired and the estrogen(E₂) and progesterone(P) concentrations in follicular fluid were determined to define the dominant follicles( DF) and subordinate follicles( SF).Granulosa cells in DF and SF were scraped,total RNA was extracted,and then libraries were constructed,and sequenced by Illumina platform.mRNA sequences were obtained by comparing the sequenced sequences with the bovine reference genome database using SOAP V2. 0 software. Differential expression of the obtained mRNA was analyzed by DESeq 2 software,and the up-regulated genes were studied by GO analysis and KEGG pathway analysis and finally real-time fluorescent quantitative PCR was used to verify the selected representative up-regulated genes.A total of 32 346 genes were obtained through sequencing,and 194 genes were significantly up-regulated in DF granulosa cells.GO analysis results showed that all the up-regulated genes were divided into 33 groups of 3 categories,and 60.6% of the genes were involved in biological process. Genes related to cellular component accounted for 21.2%,the most of which(31)were involved in cytoplasm.The genes related to molecular function accounted for 18.2%,and 18 genes were involved in metal ion binding.KEGG pathway analysis revealed a total of 4 pathways,among which the most significant gene enrichment was axon guidance pathway.Real-time fluorescent quantitative PCR results showed that the relative expression of cytochrome P450 19A1 gene(CYP19A1) in DF granulosa cells was significantly higher than that in SF(P<0.01),and the relative expressions of thioredoxin binding protein gene(TXNIP),brain expression linked X2(BEX2) and serine protease 35(PRSS35) genes in DF granulosa cells were significantly higher than those in SF(P<0.05),which was consistent with Illumina sequencing.In summary,CYP19A1,TXNIP,BEX2,and PRSS35 may promote follicular development and cause follicular ovulation in cattle.

Effect of Tetrapanax papyriferus on Lactation Performance and Endocrine Hormones Related to Lactation of Mice

LIU Lili, FENG Lili, WANG Bo

2020, 49(6): 150-156. DOI: 10.15933/j.cnki.1004-3268.2020.06.020

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In order to explore the regulation mechanism of Tetrapanax papyriferus on the lactation of mammary glands from the endocrine system,in this study,the mice were perfused with high dose 12 g(crude drug)/mL and low dose 4 g(crude drug)/mL of Tetrapanax papyriferus extracts.The effect of Tetrapanax papyriferus on milk yield of mice in lactation period was detected. The levels of PRL,E₂,P,GH in serum and mammary tissue homogenate of mice were detected by ELISA technology.The protein expression levels of β-casein,PRLR,ERα,ERβ,PR,GHR in mammary tissue were tested by Western blot technology.The results displayed that compared to the control group(the same volume of saline),high dose and low dose of Tetrapanax papyriferus extracts significantly increased the milk yield of mice in lactation period(P<0.05) and obviously promoted the expression of β-casein in mammary gland of mice(P<0.05).The levels of PRL,E₂,GH in serum and mammary tissue of mice were significantly enhanced by Tetrapanax papyriferus extracts (P<0.05).Compared with the control group,the levels of PRL,E₂,and GH in serum of mice in high dose group and low dose group respectively were significantly improved by 112.50%,129.15%,223.48% and 49.84%,68.18%,100.76%.The levels of PRL,E₂,and GH in mammary gland of mice in high dose group and low dose group respectively were obviously increased by 60.68%,269.35%,105.19% and 32.27%,129.03%,53.68%.The protein expression of PRLR,ERα,ERβ and GHR in mammary tissue were also evidently upregulated by high dose and low dose of Tetrapanax papyriferus extracts (P<0.05).However,P in serum and P,PR in mammary tissue were not significantly affected by Tetrapanax papyriferus extracts(P>0.05).This study shows that Tetrapanax papyriferus can promote the lactation of mammary gland by increasing the levels of endocrine hormones related to lactation and up regulating the expression of PRLR,ERα,ERβ and GHR in mammary gland.

Development of Cimaterol Polyclonal Antibody and Establishment of Immunological Detection Method

SI Yanfang, LI Peng, GUO Dongguang, SUN Guopeng, YUE Feng, WANG Xuannian

2020, 49(6): 157-164. DOI: 10.15933/j.cnki.1004-3268.2020.06.021

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In order to establish a specific,sensitive,fast,and accurate method for the detection of cimaterol(CIM) residues,the artificial antigen was synthesized and polyclonal antibodies were prepared by immunizing rabbits,and an immunological detection method was established. Immune antigen CIM-BSA and detection antigen CIM-OVA were synthesized after CIM was coupled to carrier proteins BSA and OVA by diazotization method.The complete antigen was identified by UV,SDS-PAGE and mass spectrometry.New Zealand rabbits were immunized with CIM-BSA to prepare anti-CIM polyclonal antibodies,and a polyclonal antibody-based indirect competitive immunoassay was established for cimaterol residual detection.The results showed that the CIM artificial antigen was successfully synthesized and possessed the molecular characteristics of the complete antigen. After immunizing rabbits,anti-CIM polyclonal antibody was prepared.The titer of anti-CIM polyclonal antibody was 1∶64 000,and IC₅₀ value was 4.26 μg/L.There was no cross reaction with β2 agonist drugs such as terbutaline,ractopamine,salbutamol,zipaterol and soon.After optimizing the conditions,an indirect competitive ELISA detection method was established based on the polyclonal antibody of CIM.The lowest detection limit of this detection method was 0.04 μg/L.The immunoassay method had a linear dependence on the antigen concentration,with R²= 0.977 1 for cimaterol concentration range of 0.1—12.8 μg/L.The fitting curve equation was y=-0.246 9x+0.662 3.The recovery rates of urine and feed samples with CIM were 96.3%—103.9% and 67.0%—105.0%.In summary,the CIM immunological detection method of indirect competitive ELISA was successfully established with good application prospects.

Impact of Red-Edge Waveband of GF6 Satellite on Classification Accuracy of Spring Crops

2020, 49(6): 165-173. DOI: 10.15933/j.cnki.1004-3268.2020.06.022

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The objective of this study is to explore the application of the red edge bands of GF6 WFV image in the identification of spring crops. Based on analysis of the spectral characteristics of single image,the identification and acreage extraction of major spring crops can be effectively achieved by random forest algorithm,taking Qixian,Henan Province as a study area,and employing basic image with 8 bands,which was collected in March 25th,2019.Combined with the ground samples and sample points data,the overall classification accuracy of four schemes, J-M distance and spectral reflectance among different training samples were calculated and analyzed.The result showed that,compared with the scheme without red-edge,the overall identification accuracy of 3 types of ground objects(winter wheat,garlic and others) with one or more red-edge was enhanced,and the separability was improved by calculating the JM distance of different features. Compared with the scheme with red-edge band 1,the overall classification accuracy of red-edge band 2 was improved by 1.98 percentage points.The overall identification accuracy of 3 types of ground objects with all red-edge bands was 86.19%,the Kappa coefficient was 0.79,while the overall identification accuracy of 3 types of ground objects without red-edge was 81.56%,and the Kappa coefficient was 0.72.By introducing all red-edge bands,the overall identification accuracy of 3 ground objects was improved by 4.63 percentage points,the separabilities of winter wheat-garlic,winter wheat-other crops and garlic-other crops were increased by 0.085 6,0.076 1 and 0.025 1 based on J-M distance,respectively.Therefore,by introducing red-edge band,the rate of wrong classification and miss classification,and “Pepper salt” effect were reduced. It could improve the overall identification accuracy of crop planting area.The result of this paper will provide a reference for the application of domesticallyproduced red-edge satellite data in agriculture.

Prediction of Fish Production and Amount of Exchanged Water of Aquaculture Pond by Principal Component Analysis and BP Neural Network Algorithm

ZHANG Junbiao, CHEN Wen, CAI Chunfang, HE Jie

2020, 49(6): 174-180. DOI: 10.15933/j.cnki.1004-3268.2020.06.023

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The fish production and exchanged water amount of aquaculture ponds are important indexes to evaluate their economic benefits and ecological effects,respectively.To explore the main factors affecting the fish production and the amount of exchanged water of conventional carp ponds,and to build forecasting models based on back propagation neural network ( BP-ANN) algorithm,51 complete managemental records about the polyculture ponds of grass carp(Ctenopharyngodon idellus),crucian carp(Carassius auratus) and common carp (Cyprinus carpio) were obtained through survey.PCA showed that, 15parameters including stocking density of grass carp and crucian carp,size of stocked crucian carp,size of harvested common carp and bighead carp(Aristichthys nobilis),production of common carp,cost of pond renting,fingerlings,feed,electric power,material for water quality regulation,costs for disease prevention and salary,water depth,occurrence of algal bloom were screened into both models of the total production of stocking fish and the amount of exchanged water in summer. The production of crucian carp and bighead carp,and oxygenation mode were also screened into the model of the total production of stocking fish.The stocking density of common carp and the stocking time of silver carp (Hypophthalmichthys molitrix)and bighead carp were also screened into the model of the amount of exchanged water in summer.45 records were randomly selected to model by BP-ANN algorithm and the other 6 records were used to predict.The results showed that the values of maximum relative error,maximum absolute error,mean relative error and mean absolute error of the model of total production of stocking fish were 8.40%,2.53 t/ha,5.81%,1.51 t/ha,respectively.The values of maximum absolute error and mean absolute error of the model of the amount of exchanged water in summer were 19.10,13.36 cm,respectively.The values of R² of the two models were 0.941 3 and 0.996 5,respectively.The values of mean square error were 0.006 5 and 0.063 3,respectively.These results demonstrated that,both models had good predictive performance,which indicated that BP-ANN algorithm was an effective means to establish models of economic benefits and ecological effects of fish ponds.

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