Journal of Henan Agricultural Sciences

Ｒesearch Progress on the Expression Technology of the Cap Protein of Porcine Circovirus Type 2 in Vitro

ZHANG Zhuqing, Lü Qizhuang, ZHUO Yanling, DENG Qixia

2019, 48(9): 1-6. DOI: 10.15933/j.cnki.1004-3268.2019.09.001

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Porcine circovirus type 2( PCV2) is the main pathogen of porcine circovirus associated diseases (PCVAD) .PCV2 infection can cause weakened immunity and immunosuppression，and thereby trigger co infection or secondary infection by a number of viruses and( or) bacteria in pigs，which brings great difficulty to the diagnosis and treatment of PCVAD.At present，vaccine immunization is the most effective measure to prevent and control PCVAD，especially the PCV2 genetic engineering vaccines that have been newly developed from the target antigen of capsid( Cap) protein． However，the acquisition of abundant， soluble and bioactive Cap proteins is crucial for developing the genetic engineering vaccine successfully.This article summarized the research progress on the expression technology of PCV2 Cap protein in vitro by using Escherichia coli，yeast，virus，bacterial vector and so on，with the purpose of providing reference for the development of rapid detection technology and genetic engineering vaccine of PCV2．

Effects of Interaction of Cutting Seedlings with Planting Density on Growth and Yield of Spring Maize

LIU Yulan, CHEN Dianyuan, ZHENG Yushi, FAN Wenzhong, LI Yongpeng, SUN Shan, WANG Tao, ZHAI Wenbo

2019, 48(9): 7-13. DOI: 10.15933/j.cnki.1004-3268.2019.09.002

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Taking Xianyu 335 as experimental material，the effects of cutting seedlings［cutting seedlings treatment( A1) ，non-cutting seedlings treatment(A2) ］and planting density［60 000(B1) ，70 000(B2) ， 80 000(B3) ，90 000(B4) and 100 000(B5) plants/ha］on the growth and yield of spring maize in Jilin Province were studied，so as to provide a new technical way for increasing spring maize yield under high density cultivation condition.The results showed that with the increase of planting density，the growth process was delayed，and the growth period was prolonged for 1—7 days; plant height and ear height increased; stem diameter，chlorophyll content，leaf area per plant and area of three ear leaves decreased; ear traits tended to deteriorate，which(except 100-grain weight) of cutting seedlings treatment were not significantly different under B1，B2 and B3 conditions，and superior to conventional planting treatment (A2B1) ; yield，harvest rate of ear decreased，and water content of grain increased on the whole.Under the same planting density，cutting seedlings delayed the growth process of spring maize，and prolonged growth period for 2—3 days; plant height，ear height，node number，leaf area per plant，area of three ear leaves and barren tip length (except A1B5 treatment) of cutting seedlings treatments decreased; chlorophyll content，number，stem diameter，ear length，ear diameter，grain number per row，100-grain weight，grain number per ear and water content of grain of cutting seedlings treatments increased; root shoot ratio of cutting seedlings treatments increased at big trumpet stage and silking stage; harvest rate of ear of cutting seedlings treatments increased; yield of cutting seedlings treatments extremely significantly increased，and the yields of A1B1，A1B2 and A1B3 treatments were extremely significantly higher than that of other treatments，which increased by 13.81%，24.13% and 21.85% compared with A2B1 treatment，respectively.To sum up，timely cutting seedlings could increase spring maize yield under the planting density of 60 000—80 000 plants/ha．

Effects of Nitrogen Fertilizer on Vascular Bundle Structure in First Internode of Super Hybrid Ｒice

WANG Liuhang, PENG Ting, XIONG Jiabao, WANG Haibin, LIU Ye, ZHANG Jing, WANG Daizhang, TENG Yongzhong, ZHAO Quanzhi

2019, 48(9): 14-22. DOI: 10.15933/j.cnki.1004-3268.2019.09.003

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In this study,filed experiment was conducted to explore the change of vascular bundle structure in first internode of super hybrid rice during grain filling stage and its relationship with yield and its components under different nitrogen(N) application rate treatments［0(N0) ，150(N10) ，210(N14) ，300 (N20) ，390(N26) ，450(N30) kg/ha］，so as to provide theoretical basis for the physiological mechanism study of increasing yield of super hybrid rice.The results showed that the application of N could change vascular bundle structure in first internode of super hybrid rice.With the increase of N application rate， the number，area and total area of small and large vascular bundles of various varieties increased first and then decreased，which were the best under the N application rate of 210 kg/ha.The number，area and total area of large and small vascular bundles in first internode of super hybrid rice were significantly negatively correlated with the number of effective panicles(except the number of large and small vascular bundles) and 1 000-grain weight，but significantly positively correlated with the grain number per panicle，seed setting rate(except the number and area of small vascular bundle) and yield(except the area of small vascular bundle) .In conclusion，reasonable application of N could affect the ability of “flow”by changing the structure of vascular bundle，which was mainly reflected in improving the grain number per panicle and seed setting rate and then increasing the yield of super hybrid rice，and the N application rate of 210 kg/ha was the best．

Path Analysis between Yield of Post-Paddy Wheat and Meteorological Factors under Different Sowing Dates in Southern Henan

ZHOU Guoqin, XIE Xudong, JIANG Mingbo, CHEN Zhenzhen, LI Gang, LI Yufeng, XIE Yingxin

2019, 48(9): 23-29. DOI: 10.15933/j.cnki.1004-3268.2019.09.004

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Aiming at high temperature stress problem at grain filling stage of post-paddy wheat in southern Henan，the winter wheat yield and its relationship with meteorological factors were studied under different sowing dates from 2015 to 2017，so as to find the suitable sowing date to avoid high temperature stress at grain filling stage.The results showed that compared with other sowing dates，the treatment with sowing date of mid October had lower mean daily temperature and mean daily relative humidity，but more sunshine and rainfall at grain filling stage; the treatment with sowing date of late October had more sunshine; the treatments with sowing date of early and mid November had higher temperature and less sunshine at grain filling stage.The grain yield and its components decreased with the postponement of sowing date，the yield of the treatment with sowing date of mid October was the highest，increased by 1.64%—129.61%，the spike number increased by 3.06%—73.25%，the 1 000-grain weight increased by 1.44%—24.98%，and the grain number per spike increased by 5.62%—32.71% compared with other treatments.The results of path analysis showed that the mean daily temperature and mean daily relative humidity had negative effect on 1 000-grain weight and grain number per spike，the sunshine hour had positive effect on 1 000-grain weight and grain number per spike from 2015 to 2017.The diurnal temperature range had positive effect on 1 000-grain weight and grain number per spike，but the rainfall had negative effect on grain number per spike from 2015 to 2016.The diurnal temperature range had no significant effect on 1 000-grain weight and grain number per spike，and the rainfall had positive effect on 1 000-grain weight and grain number per spike from 2016 to 2017.It could be seen that the main meteorological factors limiting the increase of 1 000-grain weight and grain number per spike of winter wheat were high temperature and high humidity，and the main meteorological factor that had positive effect on the 1 000-grain weight and grain number per spike of winter wheat was sunshine hour.In conclusion， the diurnal temperature range，rainfall and sunshine hour of the treatments with the sowing date of mid and late October were higher at grain filling stage，which had greater direct positive effects on 1 000-grain weight and grain number per spike，and relived the negative effect of high temperature and humidity.The temperature and humidity were high，and the sunshine hour and diurnal temperature range were low， which had higher negative effect on 1 000-grain weight and grain number per spike under the sowing date of early and mid November，so the yield significantly decreased．

Variation Analysis of Seed Quality and Antioxidant Capacity in Sesame under Drought Stress

WEI Qichao, ZHANG Haiyang, LIU Wenping, WANG Xuede, MIAO Hongmei

2019, 48(9): 30-39. DOI: 10.15933/j.cnki.1004-3268.2019.09.005

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In this article，potted simulated drought environment treatment was applied to explore the variation of the quality and antioxidant capacity of sesame seed under drought stress to reveal its response mechanism.12 sesame varieties were collected to analyse the drought effect on seed yield，seed coat color，and content of oil，protein，sesamin，sesamolin，polyphenol and antioxidant capacity.The results showed that the plant height，capsule stem length，capsule number per plant and plant yield of 12 varieties under drought stress were lower than those of the control(normal irrigation) ，of which the plant yield decreased by 2.31%—51.22%.Meanwhile，the ΔE value of seed color varied from 0.96 to 3.30，and the seed coat color got dark compared with control.The variation extents of oil content and protein content compared with control were from－0.72% to 4.03% and－5.77% to 4.12%，respectively.At the same time，the content of linoleic acid(18∶2) of 11 secame varieties seed increased by 0.44%—9.05%，and the content of oleic acid(18∶ 1) decreased by 0.16%—6.40% because of drought stress． The polyphenol content of 12 sesame varieties seed increased by－21.92%—33.00%，while FＲAP(ferrous reduction power value) increased by－24.88%—73.92% compared with control.The effects of environmental factor(i.e.,drought stress) on sesame of thousand seed weight，oil content，FＲAP value were significant，while its effects on sesamolin content and polyphenol content were evidently significant．Moreover，the significantly positive correlation between sesamolin content，polyphenol content and antioxidant capacity was found．In summary，drought stress led to lower sesame yield and deeper grain color compared with the control.Nevertheless，it was conducive to the accumulation of linoleic acid (18∶2) in sesame． Drought stress mainly affected the accumulation of sesamolin and polyphenol in sesame.So we suggest that the contents of sesamolin and polyphenol can be used as evaluation indexes for the drought resistance of sesame．

Characterization of Nitric Oxide Sensitive Mutant rsn1 in Arabidopsis

XIA Jinchan, CONG Ｒenyuan, ZHANG Xiaoli

2019, 48(9): 40-45. DOI: 10.15933/j.cnki.1004-3268.2019.09.006

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In order to investigate the molecular regulation mechanism of nitric oxide(NO) in plant，using Arabidopsis as research materials，we screened NO-sensitive mutants and isolated a mutant rsn1 (root sensitive to NO) from the EMS mutant library． The sensitivity of rsn1 mutant roots to NO and stomatal response to abscisic acid(ABA) were compared and analyzed，and the mutant genes were preliminarily located by map-based cloning.The results were as follows: the rsn1 root was more sensitive to NO stress compared with the wild type，though the germination rate of rsn1 mutant was not significantly different from that of wild type under normal growth conditions.Under 50 μmol/L SNP treatment，the relative root length of wild type( as compared with root length in normal growth) was 48.7%，whereas the relative root length of mutant was 5.3%.By analyzing the sensitivity of stomata to ABA，it was found that rsn1 was insensitive to the regulation of stomatal closure by ABA.The stomatal diameter of rsn1 mutant was 1.9 times that of wild type under 10 μmol/L ABA treatment，though the stomatal diameter of rsn1 mutant was similar to that of wild type under normal conditions，and the water loss rate of rsn1 mutant was higher than that of wild type Arabidopsis.Genetic analysis suggested that rsn1 was a single recessive mutant，and the mutation site was initially designated on chromosome 5 by map-based cloning．

Correlation Analysis between the Sugar Brix of Stalk and Agronomic Traits，Biological Yield of Sweet Sorghum in Different Growth Stages

ZHOU Yaxing, ZHOU Wei, XU Shoujun, WANG Zhenguo, LI Ziwen, CHANG Yuxin, LIU Peng, ZHANG Yongliang

2019, 48(9): 46-53. DOI: 10.15933/j.cnki.1004-3268.2019.09.007

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The aim is to explore the correlation between the sugar brix of stalk and agronomic traits， biological yield in different growth stages of sweet sorghum，and to explicit the determinants for the sugar brix formation of sweet sorghum，so as to provide reference basis for rational evaluation and utilization of existing germplasm resources and breeding varieties.The correlation between the sugar brix of stalk in the anthesis stage，grain-filling stage，early endosperm maturation，late endosperm maturation，dough stage and the plant height，rate of juice，effective sugar content and biological yield was studied using the sweet sorghum materials of sw-1，sw-2，sw-3 and sw-4.The results showed that the correlation between the sugar brix of stalk and plant height was not obvious at different growth stages.There was a significant negative correlation between the rate of juice and sugar brix in grain-filling stage.At the grain-filling stage，the sugar brix of stalk showed a significant negative correlation with effective sugar content，and an extremely significant negative correlation with biological yield.There was a positive correlation of sugar brix between anthesis stage and grain-filling stage，while there was a negative correlation of sugar brix between anthesis stage and early endosperm maturation，late endosperm and dough stage．There was a negative correlation of sugar brix between grain-filling stage and early endosperm maturation，late endosperm maturation，while there was a positive correlation of sugar brix between grain-filling stage and dough stage．Furthermore，there was a positive correlation of sugar brix between early endosperm maturation and late endosperm maturation，dough stage．The sugar brix of late endosperm maturation was positively correlated with that of dough stage.The path analysis of sugar brix of stalk between dough stage and anthesis stage，grain-filling stage，early endosperm maturation and late endosperm maturation showed that the sugar brix of stalk in the early endosperm maturation had the greatest influence on the sugar brix of stalk during the dough stage．And the path coefficient was 1.418 3.By combining the above analysis，it is concluded that when selecting high-yield，high-brix materials in sweet sorghum breeding，the parents with high sugar brix during the early endosperm maturation should be selected．

Effects of Seed Priming on Germination and Seedling Growth of Ｒadix Isatidis under Drought Stress

MENG Hengling, LU Bingyue, SU Yilan, WANG Tiantao, ZHANG Wei

2019, 48(9): 54-59. DOI: 10.15933/j.cnki.1004-3268.2019.09.008

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In order to screen out the appropriate seed initiator for Ｒadix Isatidis，four initiators including PEG-6000，CaCl₂，GA₃ and H₂O were used to treat the Ｒadix Isatidis seeds，and then the seed germination and seedling growth characteristics of Ｒadix Isatidis were investigated under 20% PEG-6000 simulated drought stress.The results showed that all these four initiators could improve seed germination rate， germination potential and germination index in different degrees under drought stress.The 18 h water priming had the best effect on seed germination，and the seed germination rate，germination potential，and germination index were increased by 33.3%，36.6% and 106.4% respectively compared with CK2(drought stress and no initiators).The follower was 25% PEG-6000 priming，and the germination rate，germination potential，germination index were 35.2%，51.2%，and 32.8% higher than CK2，respectively.The 50 mg/L GA3 priming had a significantly higher effect than other treatments on germination index，which was increased by 171.1% compared with CK2.The 1.5% CaCl₂ had lowest effect on germination rate， germination potential and germination index，which only were increased by 16.7%，19.5% and 23.9% respectively compared with CK2.In addition，after seed priming，the plant height，root length，chlorophyll content of seedlings were significantly increased，and the relative conductivity was decreased.In conclusion，the seed priming effect was in turns: 18 h water ＞25% PEG-6000 ＞50 mg/L GA₃ ＞1.5% CaCl₂ ，which suggested that 18 h water priming to treat the seeds before sowing could greatly improve the seed germination and seedling growth of Ｒadix Isatidis．

Net Carbon Sink and Their Value of Different Land-use Types in North Central Ｒegion of Henan Province

TENG Yongzhong, BAI Baoxun, CHEN Donghai, XU Tingting, SONG Changxin, WANG Weihua

2019, 48(9): 60-66. DOI: 10.15933/j.cnki.1004-3268.2019.09.009

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In order to study the net carbon sink and their value of different land-use types,for providing basis for carbon sink ecological compensation,six land-use types including typical plant species were selected in north central region of Henan Province.The net carbon sink and their value of these land-use types and plant species were evaluated using carbon tax method.The results showed that，of all the carbon input of different land-use types，greenhouse crop was the highest［2 037.88 kg/(ha·a) ］，followed by field vegetable［1 761.36 kg/(ha·a) ］，food and cash crop［1 390.92 kg/(ha·a) ］，orchard［1 226.57 kg/(ha·a) ］,urban green land［350.76 kg/(ha·a) ］，forest［198.93 kg/(ha·a) ］.Of all the carbon fixation of different land-use types,greenhouse crop was the highest［16 283.70 kg/(ha·a) ］，followed by field vegetable［11 135.38 kg/(ha·a) ］,food and cash crop［10 694.70 kg/(ha·a) ］，orchard［5 234.00 kg/(ha·a) ］，urban green land［4 783.94 kg/(ha·a) ］,forest［4 364.11 kg/(ha·a) ］.Of all the net carbon sink of different land-use types，greenhouse crop was the highest［14 245.82 kg/(ha·a) ］,followed by field vegetable［9 374.02 kg/(ha·a) ］，food and cash crop［9 303.78 kg/(ha·a) ],urban green land［4 433.18 kg/(ha·a) ］，forest［4 165.00 kg/(ha·a) ］，orchard［4 007.43 kg/(ha·a) ］.Of all the value of net carbon sink of different land-use types，greenhouse crop was the highest［13 120 Yuan/(ha·a) ］，followed by field vegetable［8 633 Yuan/(ha·a) ］,food and cash crop［8 569 Yuan/(ha·a) ］， urban green land［4 141 Yuan/(ha·a) ］,forest［3 836 Yuan/(ha·a) ］，orchard［3 691 Yuan/(ha·a) ］.The crop net carbon sink value was higher than trees，so it was suggested that the net carbon sink of crops should be incorporated into the ecological compensation system，to reduce the carbon input of crops，improve the quality of agricultural products，and give full play to the ecological function of cleaning air of crops．

Evaluation of Soil Nutrient Status and Suggestions for Fertilization in Main Peanut Production Areas of Henan Province

SHEN Yunting, SUO Yanyan, ZHANG Xiang, SI Xianzong, YU Qiong, YU Hui, SUN Yanmin, WU Jihua, CHEN Lei

2019, 48(9): 67-73. DOI: 10.15933/j.cnki.1004-3268.2019.09.010

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To fully understand the soil nutrient status in peanut production areas of Henan Province，a total of 122 soil samples were collected from four typical peanut planting areas(Zhengyang County in south ern Henan，Fangcheng County in southwestern Henan，Minquan County in eastern Henan，Qingfeng Coun ty in northern Henan) ，and their pH value and organic matter(OM) ，total nitrogen(TN) ，available phos phorus(aP) ，available potassium(aK) ，available sulfur(aS) ，available iron(aFe) and available zinc (aZn) contents were compared and evaluated.The results showed that，the average of SOM content in the Henan Province production area was 15.61 g/kg，and 92.62% of soils were at the deficiency level; the averages of TN，aK，aS contents were 1.01 g/kg，120.48 mg/kg，35.60 mg/kg，respectively，and there were 45.08%，36.07%，50.00% soils at the TN，aK，aS deficiency level，respectively; the averages of aP，aFe，aZn contents were 37.92，90.58，1.16 mg/kg，respectively，and there were 7.38%，1.64%， 0.82% soils at the aP，aFe deficiency level． For different production areas，soils in the southern and southwestern production areas of Henan were acidic and weakly acidic respectively and the soils in the northern and eastern production areas of Henan were alkaline; the OM and TN contents of four production areas were all low，and their contents in southern，eastern and southwestern production areas of Henan were higher than that of northern production area.The soil aP and aFe contents of each production area were mainly at the upper-middle level，which were higher in the southern and southwest production areas of Henan than in eastern and northern production areas． The soil aK content of each production area was mainly at the lower-middle level，which was the highest in the eastern production area of Henan and the lowest in the northern production area of Henan; the soil aS content was the highest in southern production area of Henan，but was deficient in other three production areas． The order of soil aZn contents of four pro duction areas was eastern production area＞southwestern production area＞northern production area＞southern production area.Therefore，in the peanut production area of Henan Province，people should in crease the application of organic fertilizer and potash fertilizer，strictly control the application rate of phos phate fertilizer，and carry out targeted fertilization according to different regions，add the application rate of Zn fertilizer in the southern production area of Henan，and increase the application rate of S fertilizer in the eastern，northern and southwest production areas of Henan．

Cloning and Expression Analysis of Chymotrypsin Gene PrCT1 of Pieris rapae

ZHANG Xian, LI Chaolin, ZHENG Qiaomu, YAN Zicheng, LIAO Hai, ZHOU Jiayu

2019, 48(9): 74-81. DOI: 10.15933/j.cnki.1004-3268.2019.09.011

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Chymotrypsin is the main digestive enzyme of insects and may have many other functions.In order to understand the gene expression characteristics of this protein，a chymotrypsin gene PrCT1 was cloned and analyzed based on the transcriptome data of Pieris rapae.The gene included an OＲF of 861 bp and encoded a polypeptide of 286 amino acids with a typical serine protease catalytic triplet(H57 ，D102 and S195 ) .PrCT1 was shown to be a member of chymotrypsin by analysis of the serine protease substrate-spe cific pocket.PrCT1 shared a high sequence identity( 49%) and had the closest relationship with the chy motrypsin of Danaus plexippus(GenBank accession number: OWＲ53227) .The prokaryotic recombinant vector pET28a-PrCT1 was constructed and transformed into E.coli Ｒosetta(DE3) strain to express the target protein.The results showed that PrCT1 protein existed as inclusion bodies with molecular mass of 33.26 ku.The expression of PrCT1 mＲNA in different tissues of Pieris rapae and the impacts of starvation and Cassia obtusifolia trypsin inhibitor feeding on the expression of PrCT1 mＲNA were detected by qＲT PCＲ.The results showed that PrCT1 gene was mainly expressed in the midgut of Pieris rapae，and the ex pression level was down-regulated after starvation and Cassia obtusifolia trypsin inhibitor feeding treat ments.As a result，PrCT1 might be an important enzyme taking part in food digestion in the midgut of Pieris rapae，and might be an important target of insecticide．

Cloning and Expression Analysis of PvAＲP1 Gene Encoding Auxin-regulating Protein in Common Bean

XUE Ｒenfeng, FENG Ming, ZHAO Yang, CHEN Jian, LI Tao, GE Weide

2019, 48(9): 82-89. DOI: 10.15933/j.cnki.1004-3268.2019.09.012

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Auxin-regulating protein (AＲP) is an important factor involved in the regulation of plant growth and development，and is also a protein molecule that plays an important role in plant disease resist ance． In order to understand the gene expression characteristics of this protein，the cDNA sequence enco ding the AＲP protein was cloned and analyzed by using the expression sequence tag(EST) of common bean(Phaseolus vulgaris L.) .Sequence analysis indicated that the cDNA fragment was 2 428 bp in length and named as PvAＲP1.It contained an open reading frame(OＲF) of 1 818 bp，and the accession number was MK301448.The gene encoded 605 amino acids and the predicted protein molecular weight was 68.26 ku.The encoded protein did not contain transmembrane region and signal peptide.The results of homolo gous analysis showed that the close relationship between PvAＲP1 protein and AＲP protein of Vigna angu laris reached 94%.eal time PCＲ analysis showed that PvAＲP1 gene was regulated by Fusarium oxyspo rum f.sp.phaseoli FOP-DM01 isolate.The expression of PvAＲP1 gene at 24 h after inoculation was in creased to 111 times of that at 0 h as control in the roots of the resistant material 260205 to the disease， and the expression level in 260205 roots was higher than the susceptible material BＲB130 at all time points.The expression of PvAＲP1 gene was significantly enhanced by indole-3-acetic acid(IAA) .In addi tion，the expression level in the flowers and pods was obviously higher than the roots，stems and leaves.The PvAＲP1 gene could be expressed as a 68 ku protein in E.coli BL21(DE3) ，which had the amide synthetase activity in vitro，and might regulate the active IAA level in plant cells.This study indicated that the PvAＲP1 gene probably participated in the defense response of common bean to the FOP-DM01 isolate through the IAA-mediated signaling pathway.It is speculated that the PvAＲP1 gene is related to the resist ance of fusarium wilt in common bean，and has the great significance for directing the molecular breeding of resistance in common bean．

Ｒesponse of Expression Level of Tolls and Antimicrobial Peptides Genes to Population Density in Bean Beetle

CHEN Qianquan, WANG Yeying

2019, 48(9): 90-97. DOI: 10.15933/j.cnki.1004-3268.2019.09.013

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In order to unveil the response mechanism of genes related to immunity signal transduction and antimicrobial peptide to population density，tBLASTn was used to identify genes related to Toll，IMD(Im mune deficiency) and antimicrobial peptide genes from ＲNA-sequencing data of bean beetle(Callosobru chus maculatus) .FPKM(Fragments per kilobase per million mapped reads) value was used to measure the expression level of these genes.The results showed that six Toll signaling pathway receptor genes which encoded proteins containing leucine-rich repeats domains were identified.Toll-1 had the fastest evo lutionary rate，while the evolutionary rates of Toll-3，Toll-10，Toll-7，Toll-8 and Toll-6 decreased in turn．IMD，an intracellular key signal gene，and Peptidoglycan recognition protein LC(PGＲP-LC) ，a receptor gene of IMD signaling pathway，were identified． IMD and PGＲP-LC had similar evolutionary rate to Toll 3.At expression level，Toll-1 was the lowest(FPKM=0.32) ，and was not regulated by population densi ty.The expression levels of Toll-3，Toll-6，Toll-7，Toll-8，and Toll-10 were significantly or extremely signif icantly up-regulated in high-density individuals，with Toll-7 having the highest expression level(its FP KMs in high-density individuals and low-density individuals were 127.25 and 57.92，respectively) ． Popu lation density had no significant effect on the expression level of IMD and PGＲP-LC.Seven antibacterial peptide genes were identified，including one Defensin(defensin gene) ，one Cecropin(cetanin gene) ，two Apidaecins(bee antibacterial peptide genes) ，and three Drosomycins(anti-fungal peptide genes) .Among them，Drosomycin was the most conservative one in evolution.Antibacterial peptide Apidaecin-1 had the lowest expression level(FPKM=6.58) and Defensin had the highest expression level(its FPKMs in high density individuals and low-density individuals were 1 277.23 and 683.87，respectively) .Population den sity had no significant effect on the expression level of Apidaecin-1，Cecropin and Apidaecin-2.Defensin and three Drosomycin genes were up-regulated in high density individuals.In summary，Toll-3，Toll-6， Toll-7，Toll-8，Toll-10，Defensin，Drosomycin-1，Drosomycin-2 and Drosomycin-3 were up-regulated under high population density．

Control Effect of Different Pesticides on Main Diseases and Pest Insects in Perennial Ｒice Field

YANG Shaowu, ZHANG Xiaoming, GUO Haiye, QUAN Yunchou, FU Yang, CHEN Guohua

2019, 48(9): 98-102. DOI: 10.15933/j.cnki.1004-3268.2019.09.014

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In order to clarify the effect of different pesticides on main diseases and pest insects of peren nial rice，according to the occurrence of main diseases and pest insects of perennial rice，different control schemes were formulated.Field efficacy trials were carried out in the year of 2017.The results showed that 70% imidacloprid WG and 40% virtako WG had higher control efficiency on rice planthopper than the liquid mixed with 25% actara WG and 77.5% dichlorvos EC in perennial rice field，and the control effects reached the highest level on the fifth day after spraying，which were 91.00% and 86.84%，respec tively.The field control effects of 3% carbofuran GＲ and 40% virtako WG on Cnaphalocrocis medinalis exceeded 90% and 70%，respectively，which were both significantly higher than that of 16 000 IU/mg Bacillus thuringiensis.Moreover，mixed 20% jinggangmycin SP，60% azoxystrobin WG and 70% epoxi conazole WG application had good effect on sheath blight and smut of perennial rice，and the control effi ciency was up to 83.45% and 84.45%，respectively．

Evaluation of Ｒesistance to Stripe Ｒust and Molecular Detection of Ｒesistant Genes in Wheat Germplasms at Home and Abroad

CHEN Xiangdong, WU Xiaojun, HU Tiezhu, LI Xiaohui, LI Gan, HOU Kaixin, ＲU Zhengang

2019, 48(9): 103-110. DOI: 10.15933/j.cnki.1004-3268.2019.09.015

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One hundred and twenty-four wheat germplasms were evaluated using 5 molecular markers closely linked to Yr10(Yellow rust 10) ，Yr15 and Yr24，which were resistant to stripe rust on chromosome 1BS，and disease grade was analyzed at adult stage for 4 years in field，so as to provide reference for ra tional utilization of wheat disease-resistant resources，rational distribution of disease-resistant varieties and resistance improvement of wheat varieties.The results showed that among 124 wheat germplasms，29 germ plasms displayed moderate resistance or above it，accounting for 23.39%，9 germplasms reached immune or near immune level，2 germplasms reached highly resistant level，and 18 germplasms reached moderately resistant level，accounting for 7.26% ，1.61% and 14.52% respectively; while 83 germplasms displayed susceptibility，4 germplasms reached moderately susceptible level，17 germplasms reached moderately to highly susceptible level，and 62 germplasms reached highly susceptible level，accounting for 3.23%， 13.71% and 50.00%，respectively; 12 germplasms displayed moderate susceptibility to moderate resist ance，accounting for 9.68%.The detection results of Yr genes showed that 32，28 and 80 wheat germ plasms carried Yr10，Yr15 and Yr24 genes，accounting for 25.81%，22.58% and 64.52%，respectively.Among 29 resistant germplasms，12 germplasms contained Yr10 gene，including Zhengyumai 9987，Nongda 189，Zimai No.1，Pandaola，CL0407 (Early) ，CL0438，09Zhiyin No.1，09Zhiyin No.2，CAＲMEN， MoS139，Moscow 39 and Aoayou No.1; only 3 germplasms contained Yr15 gene，including TAMOI，Pand aola and Aoayou No．1;22 germplasms contained Yr24 gene，among which 8 germplasms reached immune or near immune level，which were Jimai 22，Bainong 64，Nongda 189，Neimai 836，Yunmai 57,09Zhiyin No.1，CAＲMEN and French bermude.Yr10，Yr15 and Yr24 genes were detected in Pandaola and Aoayou No.1，while the above genes were not detected in Zhoumai 22，Bainongaikang 58，Xinmai 19，Emai No.19，HUAYUN and MＲYC，so their resistance to stripe rust might be controlled by other genes．

Analysis of Polyploidy Induction and Initial Phenotype Difference in Petunia axillaris

JIANG Hui, YUAN Xin, FENG Naixin, ZHANG Jing, LI Yanmin, FU Zhenzhu, GAO Jie, DONG Xiaoyu, WANG Huijuan, WANG Limin, ZHANG Hechen

2019, 48(9): 111-116. DOI: 10.15933/j.cnki.1004-3268.2019.09.016

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In order to obtain and identify Petunia axillaris polyploid in a short time，this study used 5% NaClO to sterilize the seeds of P.axillaris and treated them with different concentrations of colchicine，and then transferred them to MS medium for germination.Ploidy identification was performed by flow cytome ter.The optimal induction program was determined by comparing the effects of different colchicine con centrations on the induction rate.According to the difference analysis of initial phenotype，stomata and chlorophyll，the difference of morphological indexes in P.axillaris ploidy was counted.The results showed that P.axillaris tetraploid and octoploid with higher homozygous rate were obtained.While ensuring the survival rate of the mutant plants，the highest proportions of 4x(47.37%) and 8x(21.05%) were ob tained after treatment with 0.10% colchicine for 24 hours.Prolonging treatment time(36 h) could in crease the proportion of induced material with 8x ploidy( 33.33%) .The morphology，stomata and guard cells of different P.axillaris ploidy had the following rules: 4x was more rounded than 2x mutant plants in the first to second true leaves，and the morphology was similar after the third true leaves; The morphology of 8x P.axillaris was significantly different，which was characterized by short internodes，short petiole， thick and hairy leaves，dark green leaves and slow growth; The stomata and guard cell lengths of different P.axillaris ploidy nearly multiplied with the increase of ploidy，2x and 4x stomatal tissues were nearly round，but 8x stomata and guard cells did not increase by fold in the width and became elliptical．

Study on the Storage Quality of Crystal Grapes with Different Maturity

JI Ning, WANG Ｒui, HAN Zefeng, YANG Xiangqian, CAO Sen, MA Lizhi

2019, 48(9): 117-124. DOI: 10.15933/j.cnki.1004-3268.2019.09.017

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In order to explore the commercial value of crystal grapes with different harvest maturity after storage，the crystal grapes picked in three different periods were taken as experimental objects，and PE20 fresh-keeping bags were used as packaging materials stored at (1.0 ±0.3)℃ and relative humidity of (90±5) %.The results showed that stored for 60 days，the 70% maturity fruit’s rot rate(14.42%) ，threshing rate(20.26%) ，browning rate(25.16%) ，soluble solid content(12.07%) were lower than the 80% and 90% maturity． The hardness(270.19 g) was higher than the 80% and 90% maturity，and the water con tent did not differ among the maturity during the whole storage period． The 80% maturity fruit’s rot rate (20.89%) ，threshing rate( 25.06%) ，browning rate( 33.88%) were lower than those of the 90% matu rity，and vitamin C content(0.080 mg/g) was higher than 90% maturity.In summary，the 70% maturity fruit’s storability was better than the 80% and 90% maturity，but its soluble solid content was lower，re sulting in poor taste，and the 90% maturity fruit’s vitamin C content was lower than the 80% maturity.In addition，the 90% maturity fruit’s rot，browning and threshing were serious，and the commodity value was lower than that of the 80% maturity.Therefore，the comprehensive comparison showed that the crystal grape should be harvested at the 80% maturity，which would be more conducive to the commodity value after storage．

Ｒapid Analysis of Genetic Diversity of Momordica charantia Breeding Materials by Molecular Marker Technology

WANG Guoli

2019, 48(9): 125-136. DOI: 10.15933/j.cnki.1004-3268.2019.09.018

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The object of this study is to further shorten the operation process of molecular marker techno logy in bitter gourd(Momordica charantia) and maximize its application in molecular marker assisted se lection breeding.In this paper，a tubular plant DNAout kit was used to fast extract genomic DNA from bit ter gourd seeds，then the genomic DNA of 10 bitter gourd materials were amplified by PCＲ with 11 pairs of SSＲ primers and 13 pairs of SＲAP primers，and the genetic diversity of materials was also analyzed.Results indicated that the established SSＲ and SＲAP rapid amplification reaction system could obtain clear，stable and rich polymorphic bands.A total of 195 bands were amplified by 11 pairs of SSＲ primers， of which 164 bands were polymorphic，with a polymorphic band ratio of 85.7%.UPMGA cluster analysis showed that the genetic similarity coefficients of 10 materials ranged from 0.533 to 0.845，with an average of 0.690.At the threshold of 0.69，the tested materials could be divided into two groups.A total of 231 bands were amplified by 13 pairs of SＲAP primers，of which 201 bands were polymorphic，and the poly morphic band rate was 87.5%.UPMGA cluster analysis showed that the genetic similarity coefficients ranged from 0.554 to 0.775，with an average of 0.662，and all materials could be divided into two groups at the threshold of 0.66.Since both markers can obtain abundant polymorphic bands，the molecular mark er methodology can be applied to the genetic diversity analysis of bitter gourd． Moreover，the classification results by combining the two molecular marker techniques are more convincing．

AHP-based Comprehensive Evaluation of Heuchera micrantha Varieties ———Taking the Application of Green Wall in Hangzhou Area in Winter as an Example

WANG Qiaoliang, NAN Xinge, SHI Yan, WU Fan, WU Ｒenwu, YANG Fan, CHE Kanchen, BAO Zhiyi

2019, 48(9): 137-142. DOI: 10.15933/j.cnki.1004-3268.2019.09.019

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In order to screen out the varieties of Heuchera micrantha with good performance in winter green wall application，this paper proposed a comprehensive evaluation model by AHP to evaluate the green wall ap plication of 21 varieties of Heuchera micrantha collected in Hangzhou area in winter.Leaf ornamental value， winter growth status and green wall application potential were selected to define 15 evaluation indexes in 3 grades to build the model.The results showed that，the first grade(≥4.0 scores) included 5 kinds of H.mi crantha(H.micrantha Strawberry Swirls，H.micrantha Tapestry，H.micrantha Solar Power，H.micrantha Ｒio，H.micrantha Shanghai) ，6 kinds in the second grade(3.0—4.0 scores) (H.micrantha Midnight Ｒuffles，H.mi crantha Plum Pudding，H.micrantha Ｒave On，H.micrantha Peach Flambe，H.micrantha Lime Ｒuffles，H.mi crantha Marmalade) ，7 kinds in the third grade(2.0—3.0 scores) (H.micrantha Cinnabar Silver，H.micrantha Gold Zebra，H． micrantha Paris，H.micrantha Gotham，H.micrantha Solar Eclipse，H.micrantha Cracked Ice， H.micrantha Cherry Cola) and 3 kinds in the fourth grade(≤2.0 scores) (H.micrantha Hollywood，H.micran tha Sunset Ｒidge，H． micrantha Sunrise Falls) ． According to the comprehensive evaluation result and evaluation results of each level，combined with the ornamental characteristics of H.micrantha，H.micrantha Strawberry Swirls，H.micrantha Tapestry，H.micrantha Solar Power，H.micrantha Ｒio，H.micrantha Shanghai，H.micrantha Midnight Ｒuffles，H.micrantha Plum Pudding，H.micrantha Ｒave On，H.micrantha Peach Flambe，H.micrantha Lime Ｒuffles，H.micrantha Marmalade，are recommended in the application of green wall in winter．

Preparation and Immunological Characteristics of Water Adjuvant Inactivated Vaccine of Pigeon Newcastle Disease HB Strain

LI Peiyao, GAO Jingping, TIAN Yong, XU Mingju, WANG Cunlian, LI Kai, LI Jun, LAN Jinping, ZHANG Ｒuihua, XU Tong

2019, 48(9): 143-147. DOI: 10.15933/j.cnki.1004-3268.2019.09.020

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In order to effectively control the occurrence of Newcastle disease in racing pigeons in clinical practice，the isolated pigeon Newcastle disease virus Hebei strain( HB strain) was inactivated to prepare the water adjuvant vaccine，and its physical characteristics，safety，toxicity，optimal immune dose and antibody production time，immune duration and protection effects and contrast effects were tested.The results showed that the use of 0.10% formaldehyde，37 ℃，16 h inactivation could completely inactivate the virus and had the least impact on the virus titer.The water adjuvant vaccine was safe and non-toxic， and antibodies were produced at 7 d after immunization with 0.3 mL，which reached 8.1 log2 at 21 d，and reached a peak of 9.0 log2 at 28 d.The protective index was 100.0 after 30 days of immunization.Com pared with the chicken Newcastle disease vaccine(LaSota strain) ，the developed water adjuvant vaccine had a 100% protection rate，while the Chicken Newcastle disease vaccine(LaSota strain) only had 30%.It could be stored for 180 d at 4 ℃and 90 d at 20 ℃.In summary，the water adjuvant vaccine prepared according to the pigeon Newcastle disease virus HB strain could produce effective antibodies in a short pe riod of time after immunization，and the protection rate of the Newcastle disease vaccine was higher than that of the chicken Newcastle disease vaccine when racing pigeon was attacked by a strong poison．

Ｒegressive and Principal Component Analyses of Testis Weight and Secondary Sexual Characteristics at Different Weeks of Ningdu Huang Ｒoosters

ZHOU Min, ZHU Xuenong, TAN Yuwen, ZHANG Yi, WANG Yuanbin, LUO Jinsheng, ZHU Sha, YU Yan, ZHENG Shijun

2019, 48(9): 148-156. DOI: 10.15933/j.cnki.1004-3268.2019.09.021

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This experiment was conducted to study the internal correlation between testicular weight and secondary sexual characteristics at different weeks in Ningdu Huang roosters.Total testicular weight at 16 weeks of age and secondary sexual characteristics at 7 different weeks were measured.The correlation co efficients between these traits were analyzed． Moreover，regression and principal component analyses were performed among the total testicular weight at 16 weeks of age and secondary sexual characteristics at 7 different weeks.The results showed that among the characteristics measured，the variation coefficient of to tal testicular weight at 16 weeks of age was the largest(85.89%) .The variation coefficients of secondary sex characteristics such as comb length，comb height，comb thickness，length of wattle and thickness of wattle were 15.40%—37.90%，and the variation coefficients of shank were the smallest(5.13%—11.77%) .The total testicular weight at 16 weeks of age was significantly(P＜0.05) or extremely signifi cantly positivly correlated(P＜0.01) with comb，wattle and shank traits at different weeks，except for shank diameter at 16 weeks，in Ningdu Huang roosters.The correlation coefficient between total testicular weight at 16 weeks of age and the comb length at 16 weeks of age was the largest(0.688) .Stepwise re gression analysis showed that the comb length had the greatest effect on the total testicular weight at 16 weeks of age from 12 weeks，followed by the wattle length.According to the results of principal component analysis，the total testicular weight at 16 weeks of age，comb length，comb hight，comb thickness and wat tle length could be classified into the same principal component from 12 weeks． These results indicated that the comb length and the wattle length at 12 weeks of age should be considered as important indicators in the breeding for total testicular weight at 16 weeks of age in Ningdu Huang roosters．

Association between Polymorphism of Exon 1 of ESＲ1 Gene and Early Growth Performance in Quail

CAO Heng, YANG Shuai, BAI Junyan, FAN Hongdeng, PANG Youzhi, FU Xueyan, LU Xiaoning, SHI Kunpeng

2019, 48(9): 157-161. DOI: 10.15933/j.cnki.1004-3268.2019.09.022

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In order to investigate the effect of polymorphism of estrogen receptor gene 1(ESＲ1) on early growth performance of quails，three quail populations(Chinese yellow quail，Korean quail and Beijing white quail) were tested by PCＲ-ＲFLP，and the association between ESＲ1 gene exon 1 and early growth performance of quails was analyzed.The results showed that three genotypes TT，CC and CT were detected in exon 1 of ESＲ1 gene in three quail populations.The CC genotype frequencies of exon 1 of ESＲ1 gene were 0.515，0.600 and 0.723，TT genotype frequencies were 0.088，0.014 and 0.015，and CT genotype frequencies were 0.397，0.386 and 0.262 respectively in Chinese yellow quails，Beijing white quails and Korean quails.The correlation analysis showed that exon 1 of ESＲ1 gene was correlated with tibia length， sternum length，body length and tibia circumference of Korean quails(P＜0.05) .Exon 1 of ESＲ1 gene was correlated with body weight，tibia length，chest depth，body length and sternum length of Chinese yellow quail(P＜0.05) .Exon 1 of ESＲ1 gene was also correlated with body mass of Beijing white quail (P＜0.05) .These results suggest that exon 1 of ESＲ1 gene can be used as a candidate gene for marker assisted selection of early growth performance of egg quails．

Toxic Effects of Cyhalofop-butyl to Misgurnus anguillicaudatus

SHANG Taiyu, SHI Chunyu, QIN Zhaohui, PENG Mengyuan, JIA Boshuang, XIA Xiaohua

2019, 48(9): 162-167. DOI: 10.15933/j.cnki.1004-3268.2019.09.023

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In order to estimate the herbicide toxicity of cyhalofop-butyl to aquatic organisms，with Misgur nus anguillicaudatus as the experimental object，toxic effects of cyhalofop-butyl on Misgurnus anguillicau datus including acute toxicity，physicological toxicity and histomorphological toxicity were studied.The re sults of acute toxicity test showed that the half lethal concentration(LC₅₀) of cyhalofop-butyl to Misgurnus anguillicaudatus in different periods( 24 h，48 h，72 h and 96 h) were 7.254 4 mg/L，7.102 3 mg/L and 6.809 3 mg/L and 6.623 7 mg/L，respectively.The safety concentration(SC) was 2.042 3 mg/L.Through physiological toxicity experiments，the activities of SOD and CAT in Misgurnus anguillicaudatus’s liver increased obviously，and the maximum value appeared in the high concentration treatment group (1.324 8 mg/L) on the first day. The maximum activity value of GSH-Px appeared in the high concen tration treatment group(1.324 8 mg/L) on the third day.Meanwhile，the maximum value of the expres sions of SOD and CAT appeared in the low concentration treatment group( 0.662 4 mg/L) on the seventh day.The expression of GSH-Px reached the maximum on the third day.And these had significant differ ences compared with the control(0 mg/L).Overall，with the extension of exposure time and the concen tration increasing，the activities and gene expressions of SOD，CAT and GSH-Px initially increased and then decreased in the liver of Misgurnus anguillicaudatus.In the histomorphological toxicity test，the wid ened interstitial space could be observed in hepatocytes on the fifth day，while hepatocytes were swelling and more vacuolating on the seventh day in the low concentration treatment group(0.662 4 mg/L) .More over，vacuolation was observed on the first day in the high concentration treatment group(1.324 8 mg/L). With the prolongation of exposure time，hepatocytes swelling and vacuolation deepened，and karyopyknosis could be observed.The experiment results showed that the cyhalofop-butyl had certain toxicity and damage effect on antioxidant enzyme system and histocyte of Misgurnus anguillicaudatus’liver，so it should be used reasonably and scientifically．

Ananlysis of Agricultural Ecological Security Knowledge Map Based on Web of Science

SUN Yan, ZHANG Huifang, DUAN Junzhi, QI Hongzhi, YANG Cuiping, FAN Wenjie, YAN Zhaoling, CHEN Haiyan, LIU Yuxia, ZHUO Wenfei

2019, 48(9): 168-176. DOI: 10.15933/j.cnki.1004-3268.2019.09.024

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For the reveal of historic dynamic of agricultural ecological security area，to bring reference for theoretical and practical of it in China，we used Web of Science as data resources，CiteSpace as tool，and analyzed knowledge map of agricultural ecological security area in 2008—2018.The results showed that， agricultural ecological security area development was divided into 3 stages: basic stage(2008—2011) ， rapid growth stage( 2012—2014) ，new rapid growth stage(2015—2018).Different stage had different re search emphasis.In a total，agricultural ecological security research focused on food security，and extended to evaluation，spatial pattern，ecological risk assessment aspect，etc.TILMAN D，GODFＲAY H C J，FO LEY J A，ALTIEＲI M A，LOBELL D B，FOLKE C，PＲETTY J were the leading researchers of this area， and contributed a lot in this area.Main journals that published more papers in agricultural ecological se curity area were Sustainability，Food Security，Land Use Policy，PloS One，etc.Chinese Academy of Sci ences showed the absolute centrality in agricultural ecological security area. Papers with the foundation of National Natural Science Foundation of China were the most，whose percentage in total searched papers was 12.9%，and ranked No.1.In conclusion，agricultural ecological security research should take interna tional development tendency as reference，introduce technologies and ideas of modern agriculture and mathematical model，to refine，precise and advance agricultural ecological security research in China.We should follow the leading researchers，grasp leading edge，distribute new hot issue.Further we should ex plode open co-operation，for the purpose of consolidating and strengthening the academic status of China in agricultural ecological security area．

Application of Moisure Changing Model of Tobacco Leaves during Bulk Curing Process

LI Shengdong, WANG Tao, GAO Yabei, XIE Yan, ZHANG Baoquan, MA Liujun, SONG Zhaopeng, LOU Xiaoping

2019, 48(9): 177-184. DOI: 10.15933/j.cnki.1004-3268.2019.09.025

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In order to perfect and improve the moisture changing model curing system of tobacco leaves and provide some relevant data support，the flue-cured tobacco leaf comprehensive benefit and quality by tobacco leaf moisture changing model( Wang and singh model) curing and conventional curing were studied.Tobacco variety Yunyan 87 in Ji’an，Jiangxi was chosen as the experimental material.Combined with the value of water loss at 7 stages during flue curing process，using Wang and singh model for mathematical formula calculation and derivation，the economic benefit，grade structure，chemical composition and sensory quality of flue-cured tobacco leaves were analyzed.Compared with the conventional curing，in terms of economic benefit，model curing could increase the yield of tobacco leaves by 60 kg/ha，increase the output value by 2 500 Yuan/ha，and increase the average price by 0.48 Yuan/kg.In terms of grade structure，model curing could significantly increase the proportion of orange yellow leaf by 5.55 percentage points，increase the proportion of first class leaf by 4.52 percentage points，and reduce the proportion of inferior leaf by 0.41 percentage points.In terms of chemical composition，model curing could significantly increase the total sugar，reducing sugar content and the value of reducing sugar/nicotine of flue-cured tobacco leaves，the second-level of orange lower leaf(X2F) ，the third-level of orange middle leaf(C3F) and the second-level of orange upper leaf(B2F) total sugar content increased by 6.06，2.63，2.48 percentage points，respectively; reducing sugar content increased by 3.01，2.82，2.55 percentage points，respectively; reducing sugar/nicotine value increased by 3.30，2.30，0.93 points.And the chemical composition coordination of C3F，B2F increased by 2.03，4.66 points，respectively.In terms of sensory quality，model curing could improve the score of fragrance quality，fragrance quantity， concentration and strength.Model curing could significantly improve the X2F，C3F，B2F total scores by 1.63，2.25，0.59 points，respectively，while sensory quality coordination could be improved by 0.40，0.22 and 0.19 points，respectively.The Wang and singh model curing method obtains a good comprehensive quality evaluation of flue-cured tobacco leaves.The model can be used to realize accurate tobacco leaves curing under the premise of limited water loss and curing time．

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