Loading...

Table of Content

    16 May 2018, Volume 51 Issue 10
    CONTENTS
    Scientia Agricultura Sinica. 2018, 51(10):  0-0.  doi:10.3864/j.issn.0578-1752.nykx-201851-1000
    Abstract ( 117 )   PDF (322KB) ( 165 )   Save
    Related Articles | Metrics
    CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS
    TaDRO, A Gene Associated with Wheat Root Architectures, Its Global Distribution and Evolution in Breeding
    WeiJun ZHANG, Tian LI, Lin QIN, Jing ZHAO, JunJie ZHAO, Hong LIU, Jian HOU, ChenYang HAO, DongSheng CHEN, YiQin WEI, RuiLian JIN, XueYong ZHANG
    Scientia Agricultura Sinica. 2018, 51(10):  1815-1829.  doi:10.3864/j.issn.0578-1752.2018.10.001
    Abstract ( 470 )   HTML ( 38 )   PDF (3498KB) ( 253 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 Root improvement is one of the key factors to improve stress resistance and yield of wheat. Root architecture related genes, homoeologous TaDRO, are cloned from cultivars with different root phenotypes. Molecular markers are developed to detect its relationship with important agronomic traits of wheat. which could provide technical support for wheat improvement. 【Method】 Polymorphic sites of TaDRO-A, -B and -D were detected in 21 common wheat accessions with high diversity. Physical locations of three homoeologues were determined based on the newest genome sequence of Chinese Spring. Molecular markers were developed according to the polymorphic sites at TaDRO-5A and -5B. Association analysis between genotypes and phenotypic traits were carried out in a natural population of 323 accessions. 【Result】 The three homoeologous genes of TaDRO-A, -B and -D were cloned. TaDRO-A, -B and -D were located on chromosomes 5A (426.15 Mb), 5B (381.00 Mb) and 5D (327.60 Mb), respectively. Three SNPs were detected at TaDRO-A among 21 accessions and two haplotypes were formed, Hap-5A-A and Hap-5A-C. A molecular marker, TaDRO-5A-KASP, was developed based on the SNP located at position of -2271 bp in the promoter region. Thirteen SNPs and one InDel in the promoter region, four SNPs in the coding region were detected at TaDRO-5B, formed two haplotypes, Hap-5B-Ⅰ and Hap-5B-Ⅱ. The marker TaDRO-5B-InDel was developed based on the Indel at position of -300 bp. Association analysis showed that haplotypes of TaDRO-5A were significantly correlated with plant height (PH), thousand kernel weight (TKW) and root growth angle (RGA). The genetic effects on Hap-5A-A showed RGA and TKW increasing, and root depth and PH decreasing, while those on Hap-5A-C exhibited the opposite effect. The effects of Hap-5B-Ⅰ exhibited root depth and PH increasing, and TKW decreasing, while those of Hap-5B-Ⅱ were the opposite. Hap-5A-C and Hap-5B-Ⅰ were favored haplotypes in landraces while they were non-favored ones in modern cultivars. The frequencies of Hap-5A-C and Hap-5B-Ⅰ in arid and semi-arid areas were higher than those in wet areas in China. Hap-5A-C was favored haplotype in dry regions worldwide. Frequencies of Hap-5A-A and Hap-5B-Ⅱ were increasing in breeding process, respectively.【Conclusion】 Hap-5A-C and Hap-5B-Ⅰ are associated with deeper root, higher PH and lower TKW, whereas Hap-5A-A and Hap-5B-Ⅱ behave the opposite. Wheat cultivars showed root depth and plant height decreasing in breeding process due to development of irrigation system and nitrogen industries. The developed molecular markers might be used to select the ideotypes of cultivars for higher efficiency use of water and nitrogen.

    Rice Overexpression of Millet SiANT1 Gene Increases Salt Tolerance
    Lei NING, ShuGuang WANG, PengJu JU, XingXuan BAI, LinHao GE, Xin QI, QiYan JIANG, XianJun SUN, Ming CHEN, DaiZhen SUN
    Scientia Agricultura Sinica. 2018, 51(10):  1830-1841.  doi:10.3864/j.issn.0578-1752.2018.10.002
    Abstract ( 365 )   HTML ( 13 )   PDF (3336KB) ( 464 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 High salt stress is one of the main factors affecting crop yield. Setaria italica L. is characterized by its strong tolerance to stress. Therefore, screening salt tolerant related genes from millet is important for salt stress tolerant improvement of crops through genetic transformation.【Method】 The expression profile of AP2/ERF like gene SiANT1 were analyzed in Yugu 1 at seedling stage under high salt, low nitrogen and PEG simulated drought conditions. Millet gene SiANT1 was transformed into rice Kitaake by agrobacterium-mediated transformation method. overexpressing SiANT1 rice seeds and wild type Kitaake were digested with 0.9% saline and tap water, respectively. The phenotypes of germinating stage and germination rate were observed. Rice seedlings were treated with 0.9% NaCl in Hogland nutrient solution for 10 days and then dried at 65℃ for 48 hours dry weights of all the rice overexpressing SiANT1 and wild type (WT) plants were analyzed. At the same time, phenotype of all plants was analyzed in the field and all plants were irrigated once with 0.9% saline before rice pregnancy. Under the similar salt treatment condition, single seed grain weigh plant height and tiller number per plant of the overexpressing SiANT1 rice lines and wild type were also analyzed. The real time-PCR method was used to verify the expression of SiANT1 and its possible downstream genes in transgenic rice lines.【Result】 Setaria italica L. SiANT1 protein (XP004985124.1, SiANT1) belongs to the AP2 subfamily and has a close genetic relationship with sorghum (XP021318293.1, SbANT1) and maize (XP008658933.1, ZmAP2).The gene of SiANT1 was induced by high salt stress (100 mmol·L-1 NaCl) in Yugu 1. The SiANT1 was ligated to LP047 1118-Bar-ubi-EDLL vector, Agrobacterium transformation method was used to transfer SiANT1 into rice genome, positive T0 plants were screened, then breeded two generations and got T3 seeds. Under water immersion, the germination rate of SiANT1 overexpression rice was similar to that of wild type. Germination rate was more than 90%. Under 0.9% saline immersion, the overexpression of SiANT1 was significantly inhibited. Compared with the wild type, over-expression of SiANT1 rice seeds dew white was delayed, but finally(treatmented for 120h) germination rate was more than 80% .After treatment with 0.9% NaCl at seedling stage in greenhouse, the dry weight of single plant overexpressing SiANT1 increased 14.11%-37.42% more than that of WT, which had significant difference at the 1% level. The grain weight of transgenic rice overexpression SiANT1 in field increased 56.12%-76.58% higher than that of WT, which had significant difference at the 5% level, the number of tillers per plant and plant height increased, but there was no significant difference with the wild-type. Semi-quantitative RT-PCR analysis showed that all three rice lines overexpressing SiANT1 expressed SiANT1 at the RNA level. Real time-PCR results showed that the relative expression level of SiANT1 in three rice lines overexpressing SiANT1 was different but significantly increased compared with that of the recipient, and the relative expression levels of endogenous salt-tolerance-related genes OsSOS1 and OsZFP182 were 1.1-1.7 fold and 1.6-2.3 fold respectively higher than that of the recipient.【Conclusion】 It has been confirmed that OsSOS1 and OsZFP182 are salt tolerance related genes. SiANT1 gene has certain salt tolerance. Overexpression of SiANT1 in rice may increase salt tolerance by increasing the expression of downstream genes OsSOS1 and OsZFP182.

    TILLAGE & CULTIVATION·PHYSIOLOGY & BIOCHEMISTRY·AGRICULTURE INFORMATION TECHNOLOGY
    Grain Mechanical Harvesting Technology Promotes the Transformation of Maize Production Mode
    ShaoKun LI, KeRu WANG, RuiZhi XIE, Bo MING
    Scientia Agricultura Sinica. 2018, 51(10):  1842-1844.  doi:10.3864/j.issn.0578-1752.2018.10.003
    Abstract ( 355 )   HTML ( 34 )   PDF (232KB) ( 409 )   Save
    References | Related Articles | Metrics
    Research Progress of Maize Lodging During Late Stage
    Jun XUE, KeRu WANG, RuiZhi XIE, Ling GOU, WangFeng ZHANG, Bo MING, Peng HOU, ShaoKun LI
    Scientia Agricultura Sinica. 2018, 51(10):  1845-1854.  doi:10.3864/j.issn.0578-1752.2018.10.004
    Abstract ( 592 )   HTML ( 20 )   PDF (1078KB) ( 923 )   Save
    Figures and Tables | References | Related Articles | Metrics

    Lodging is a common problem during maize production. Maize harvested at physiological maturity stage in traditional maize production. Previous researches about maize lodging also focused on stalk development process and some stage before the physiological maturation. There was less lodging research after physiological maturity. Mechanical grain harvest usually was carried out two to four weeks after the physiological maturity. Lodging made mechanical grain harvest become difficulty and increased harvest costs, and it also increased grain losses and decreased the grain quality. Maize lodging was a limiting factor in application of mechanical grain harvest technology. This paper reviewed the physiological changes in the maize senescence process and the influences factors on maize lodging, and proposed the measures and suggestions about improving maize lodging resistance during mid and late stage. Analysis showed that, natural senescence of maize from physiological maturity to harvest decreased activity of leaves, stalk and root, and made all of stalk moisture, water-soluble carbohydrate content, hemicellulose content and total structural carbohydrate content decline. These resulted in cell wall thinning, cell gap widening, and stalk strength and disease resistance decrease. The enzymatic activity of PAL, POD and PPO was low, which made the disease resistance decline. The stalk rot pathogen produced cell wall degrading enzymes to decompose cellulose and collapse the cells in stalk. The fungal hyphae flew through the plasma membrane and entered into epidermis cell, cortex and vascular bundle tissue. The stalk rot then increased the speed of stalk dehydration and drying shrinkage, and degraded the pith tissue. Maize stalk became hollow, softening and rot. This decreased stalk quality. Higher grain and full mechanization technologies required high population and delayed harvest to grain dehydration in the field. These accelerated and extension the aging process of maize, further decreased the stalk quality and disease resistance, and increased the lodging risk of maize after physiological maturity. In order to resolve the lodging problem and to accelerate development and promotion of maize mechanical grain harvest technology in China, we suggested that: (1) Theoretical research of maize lodging during late stage should be strengthened; (2) Germplasm creation should be strengthened to breed new maize cultivars with early-maturity, density-tolerant, high rate of grain dehydration, and strong stress resistance; (3) Common methods should be developed to obtain high-quality maize populations and reduce lodging during late stage including constructing a high-quality plow layer, suitable cultivars for grain mechanical harvest, rational close planting, scientific irrigation and fertilization, application of plant growth regulators, and insect-disease prevention; (4) Integration countermeasures should also be established according to ecological conditions in different maize regions.

    Differences of Ear Characters in Maize and Their Effects on Grain Dehydration
    LuLu LI, Bo MING, RuiZhi XIE, KeRu WANG, Peng HOU, ShaoKun LI
    Scientia Agricultura Sinica. 2018, 51(10):  1855-1867.  doi:10.3864/j.issn.0578-1752.2018.10.005
    Abstract ( 392 )   HTML ( 17 )   PDF (936KB) ( 622 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 The high grain dehydration rate and the low grain moisture content at harvest are two ear characters, which can be established for maize mechanical grain harvesting. Ear characters are decided by genes and have close relationship with grain dehydration. This internal relationship and the key ear characters that can characterize the traits of grain dehydration remain unknown, which are of great significance for breeding and screening of suitable varieties. 【Method】 The report researched on a total of 22 main cultivars of summer maize in HuangHuaiHai plain, and their ear characters were divided into 41 parameters covering bract, grain, cob and ear-pedicel. In 2015 and 2016, these parameters were measured and were used in correlation analysis with five grain parameters describing the dehydration rate, including the grain dehydration rate before physiological maturity (GDRbpm), the grain dehydration rate after physiological maturity (GDRapm), the total grain dehydration rate (GTDR), the grain moisture content at physiological maturity (GMCpm) and at harvest (GMCh). 【Result】 These 41 parameters were significantly different between the 22 cultivars, and there were some parameters significantly linked to grain dehydration. The bract length had a significantly negative relationship with GDRapm and a significantly positive relationship with GMCh. The value of “bract length/ear length” had a significantly negative correlation with GDRapm. The ear angle was positively correlated to GTDR at a significant level. The cob moisture content at physiological maturity had significantly positive relationships with GMCpm and GMCh. The grain number per ear was positively correlated to GDRbpm and GTDR at a significant level. The value of “ear length/grain number per row” had significantly positive relationships with GDRbpm, GDRapm and GTDR and had a significantly negative relationship with GMCh. The 100-grain dry weight at physiological maturity was negatively correlated to the grain moisture content at the significant level. There were no significant correlations between the other ear parameters and the five grain dehydration parameters.【Conclusion】 The current cultivars had different ear characters in HuangHuaiHai plain. These parameters contributed to grain dehydration rate, including the shorter bract, the lower moisture content of cob at physiological maturity, the larger ear angle, the fewer grain number per ear, and the smaller grain, which could be used in breeding and screening of mechanical grain harvest cultivars.

    Maize Cob Mechanical Strength and Its Influence on Kernel Broken Rate
    Jun XUE, LuLu LI, WanXu ZHANG, Qun WANG, RuiZhi XIE, KeRu WANG, Bo MING, Peng HOU, ShaoKun LI
    Scientia Agricultura Sinica. 2018, 51(10):  1868-1877.  doi:10.3864/j.issn.0578-1752.2018.10.006
    Abstract ( 289 )   HTML ( 12 )   PDF (594KB) ( 315 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 The objectives of this study were to investigate the variation of maize cob mechanical strength and its influence on kernel broken rate to provide a theoretical basis for improving maize harvest quality. 【Method】 Maize cultivars selection experiments were conducted in large area field. The same combine harvester and operating personnel were used in different harvest date to study the variation regularity and influence factors of maize cob mechanical strength during late growth period, to analyze the relationship between cob mechanical strength and kernel broken rate. The cob morphology, moisture content, dry matter accumulation, mechanical strength characters, kernel moisture content, and broken rate were investigated in this research. 【Result】 The results showed that, as harvest date delayed, maize kernel and cob moisture content decreased, and the bending strength of 8 cm and whole cob, and kernel broken rate showed a trend of decrease first and then increase. When kernel moisture content was below 20.1%, kernel broken rate increased exponentially with the increase in cob strength. When kernel moisture content was higher than 20.1%, kernel broken rate increased exponentially with the increase in bending strength of whole cob, and there was no significant relationship between kernel broken rate and bending strength of 8 cm cob. The bending strength was significantly and negatively correlated with moisture content of cob. In contrast, bending strength were significantly and positively correlated with penetration strength, dry weight, dry weight per unit length, and dry weight per unit volume. Path analysis showed that dry weight per unit length had the greatest effect on bending strength of cob. 【Conclusion】 Cob mechanical strength is one of the important factors affecting kernel broken rate in maize grain harvest. The dry matter accumulation and moisture content are important factors affecting the mechanical strength of the cob during late stage.

    Study on Grain Dehydration Characters of Summer Maize and Its Relationship with Grain Filling
    LuLu LI, Bo MING, Shang GAO, RuiZhi XIE, Peng HOU, KeRu WANG, ShaoKun LI
    Scientia Agricultura Sinica. 2018, 51(10):  1878-1889.  doi:10.3864/j.issn.0578-1752.2018.10.007
    Abstract ( 530 )   HTML ( 30 )   PDF (828KB) ( 925 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 Nowadays, the higher grain moisture content at harvest limits the popularization and application of the mechanical grain harvesting technology. Maize grain filling process is accompanied by grain dehydration process after pollination, however, the relationship between these two processes remains a challenge. We used different maize cultivars to study the characters of the two processes and the relationships between them, which provided support for breeding and promotion of the harvesting technology. 【Method】 Field experiments were conducted in Xinxiang, Henan in 2015 and 2016. A total of 22 cultivars were studied and the controlled pollination was applied in every cultivar. In 2015, the sampling time was from the 26th day after pollination to November 14th. In 2016, the sampling time was from 11th day after pollination to October 17th. We measured dynamic changes of grain moisture content (MC), moisture (M), dry weight (DW) and fresh weight (FW) before and after physiological maturity to establish the relationships between these indexes and the accumulated temperature after pollination (T) by equations. Based on these equations, the grain dehydration process and the filling process were clarified. Then, we developed the relationship between these two processes by the correlation analysis. 【Result】 Results showed that T had the significant non-linear relationships with MC, M, DW and FW. Among them, the relationship between MC and T of 22 maize cultivars could be described by the Logistic Power regression model. The MC dropped to 28% when the T reached average 1 357°C·d, changing from 1 126 °C·d to 1 646 °C·d between cultivars. The average T was 1 480°C·d for 25% MC, changing from 1 218 °C·d to 1 810 °C·d. Dynamic change of MC could be divided into two stages based on the changes of DW and M. The first stage was from the start of grain growth to the end of linear filling process, in which the decreasing MC was mainly decided by the fast dry matter accumulation. The second stage followed the former ending to the harvest time, in which the decreasing MC was owned to the decreasing M. The correlation analysis showed that there was a significant negative correlation between the MC at physiological maturity and the filling days, and the T from pollination to physiological maturity in 2015 while the relationship was not significant in 2016. There was no significant relationship between the filling rate and the grain dehydration rate before physiological maturity, similar to the grain dehydration rate after physiological maturity and the total dehydration rate. 【Conclusion】 Our study found that the Logistic Power regression model had a good predictive stability to establish the relationship between MC and T. We proposed that MC was decided by the grain filling rate and the grain moisture loss rate respectively at different stages. Thus, breeders should not only pay attention to grain filling characters and maturity time, but also concern about the grain dehydration characters when evaluate suitable cultivars for the harvesting technology.

    Analysis of Sowing and Harvesting Allocation of Maize Based on Cultivar Maturity and Grain Dehydration Characteristics
    WanXu ZHANG, Bo MING, KeRu WANG, ChaoWei LIU, Peng HOU, JiangLu CHEN, GuoQiang ZHANG, JingJing YANG, ShuLing CHE, RuiZhi XIE, ShaoKun LI
    Scientia Agricultura Sinica. 2018, 51(10):  1890-1898.  doi:10.3864/j.issn.0578-1752.2018.10.008
    Abstract ( 367 )   HTML ( 26 )   PDF (451KB) ( 581 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 Yield and production efficiency are two equally important things under the condition of large-scale production. The sowing time and the harvesting time can be prolonged by various combinations of sowing date and different maturity cultivars, thus improving the utilization efficiency of combine machine and the maize production efficiency.【Method】 In this study, three maize cultivars, including KWS9384, Xinyu77 and M751, with different growth stages were selected to monitor the dynamic process of grain moisture content from 2015 to 2017. The predictive relationship model between the grain moisture content and the accumulated temperature (> 0°C) after pollination was established to analyze the key growth nodes of different combinations based on the local meteorological data.【Result】 The results showed that there were significant differences of grain yield and suitable sowing date between cultivars. The early maturity cultivar KWS9384 had a longer time of sowing and harvesting but a lower yield compared with the late maturity cultivars. The late maturity cultivars Xinyu77 and M751 both had higher yields but they needed more time to finish physiological maturity and to dry down grain to meet grain mechanical harvest. The combination plans of late maturity cultivar/early sowing or early maturity cultivar/late sowing could be used to coordinate the relationship between yield and grain moisture content, thus extending the sowing time and the grain harvesting time.【Conclusion】 This paper studied on the suitable sowing time and harvesting time of different maturity cultivars and gave the combination principle of cultivar and sowing date under the background of high yield and high efficiency production. The principle could maximize the utilization efficiency and benefit of combine machine under the specific ecology and production condition. This study provided the new information regarding the relevant researches and application of the maize grain mechanical harvesting technology.

    PLANT PROTECTION
    Influence of Temperature and Polyamines on Occurrence of Citrus Canker Disease and Underlying Mechanisms
    Feng YANG, ChuanWu CHEN, QiJun FAN, ChunMei SHI, ZongZhou XIE, DaYong GUO, JiHong LIU
    Scientia Agricultura Sinica. 2018, 51(10):  1899-1907.  doi:10.3864/j.issn.0578-1752.2018.10.009
    Abstract ( 384 )   HTML ( 4 )   PDF (1019KB) ( 188 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 Canker disease is one of the most devastating diseases that cause serious damages to citrus. It is more likely to occur under high temperature. The objective of this study is to elucidate the mechanism underlying the disease incidence at high temperatures, reveal its metabolic changes, and to provide important theoretical guidance for controlling the disease using certain chemicals.【Method】 Sweet orange (Citrus sinensis), which is sensitive to canker disease, was used as the experimental material. The sweet orange plants were pre-cultured for 3 d at either 21℃ or 30℃ prior to inoculation with Xanthomonas citri subsp. citri (Xcc), followed by evaluation of disease incidence. Expression of four defense-related genes, including AOS (allene oxide synthase), CHI (chitinase), GPX (glutathione peroxidase) and PR4A (pathogenesis-related protein 4A), in the plants pre-cultured at the two temperatures was assessed by semi-quantitative RT-PCR. Meanwhile, endogenous polyamines (putrescine, spermidine and spermine) in the plants pre-cultured at the two temperatures were also analyzed by HPLC. In addition, sweet orange plants were treated with exogenous spermidine (0.4 mmol·L-1), using water treatment as a control, before Xcc inoculation. Disease incidence and index of plants treated with either spermidine or water were compared, while endogenous polyamine contents and expression levels of defense-related genes (AOS, CHI, GPX and PR4A) in response to spermidine or water treatment were assessed. 【Result】 After inoculation with Xcc, it was found that plants pre-cultured at 21℃ exhibited a lower cankder disease incidence at the early stage when compared with the plants pre-cultured at 30℃. On the 10th day, the incidence of the two treatments was similar. HPLC analysis showed that content of the three free polyamines (putrescine, spermidine and spermine) in plants pre-cultured at 21℃ was significantly higher than that in the plants pre-cultured at 30℃. In addition, RT-PCR analysis indicated that the transcript level of three defense-related genes, CHI, GPX and PR4A, in plants kept at 21℃ was higher than that from 30℃, while there was no significant difference in AOS expression between the two groups. Exogenous application of spermidine remarkably enhanced levels of endogenous putresicne and spermidine, reduced disease incidence and index in comparison with water treatment. Spermidine treatment reduced the disease incidence by 45% and in comparison with the control after 14 days of inoculation. In addition, the disease index of the spermidine-treated samples was 4.8 lower than that of the control. Meanwhile, the phenotype indicated that the control displayed more serious symptom than that of spermidine treatment. Moreover, spermidine treatment could up-regualte mRNA abundances of all four defense-realted genes, including AOS, CHI, GPX and PR4A. 【Conclusion】 Sweet orange displayed susceptibility to citrus canker at high temperature, and the potential mechanisms underlying this phenomenon may be ascribed to inhibition of defense-related genes and suppression of polyamine biosynthesis. Exogenous polyamine treatment conferred enhanced tolerance to citrus canker by upregulating defense-related genes and triggering disease resistance response. Taken together, high temperature is one of the environmental factors accounting for outbreak of citrus canker disease, and polyamines are conducive for improving tolerance to citrus canker disease.

    Antimicrobial Effect and Components Analysis of Volatile Organic Compounds from Bacillus pumilus AR03
    Jing WANG, JianMin CAO, DeXin CHEN, Jun QIU, XiaoQiang WANG, Chao FENG, WenJing WANG
    Scientia Agricultura Sinica. 2018, 51(10):  1908-1919.  doi:10.3864/j.issn.0578-1752.2018.10.010
    Abstract ( 466 )   HTML ( 8 )   PDF (1862KB) ( 484 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 The objective of this study is to determine the antimicrobial activity of volatile organic compounds (VOCs), produced from tobacco rhizosphere soil Bacillus pumilus AR03 strain, and to analyze its main components.【Method】 The antifungal effect of VOCs on the colony, mycelium growth and spore germination of Phytophthora parasitica var. nicotianae and Alternaria alternata was determined by a double Petri dish assay and cavity slide method. The control effect of VOCs on tobacco black shank and brown spot by leaf inoculation was determined in vitro. VOCs were collected by head-space solid phase microextraction (HS-SPME) and identified by gas chromatography-mass spectrometry (GC-MS). Retention index (RI) and internal standard (IS) 1-pentadecence were used for qualitative and quantitative analysis.【Result】 VOCs released from B. pumilus AR03 strain had certain inhibitory effect on the two target pathogens, which showed that the mycelium of P. parasitica grew slowly and thinned, branches increased and twisted, and broke. The mycelium of A. alternata was deformed and no conidial pedicle was produced on the mycelium. Most of the inclusions gathered together and caused the mycelium to dry and constricted. Growth of exposed fungus colonies was inhibited by VOCs, the inhibition rates of VOCs were 56.21% and 59.23%, 64.75% and 59.86%, 66.13% and 61.10%, 67.04% and 70.00%, respectively, against P. parasitica and A. alternata cultured in sealed plates for 2, 4, 6 and 8 d. When the zoospores of P. parasitica and ascospores of A. alternata exposed to these volatile components for 6 h, the germination was delayed and the growth was slow. The number of sporocyst produced by P. parasitica obviously reduced. Most conidiophores of A. alternata expanded abnormally as cystic structure, indicating the fungicidal nature of the volatiles. Moreover, VOCs could significantly inhibit the disease severities of tobacco black shank and brown spot on leaves tests. Direct fumigation for 40 h and 80 h, black shank disease incidence was 92.50% on control and 70.83% on leaves treated by VOCs, the inhibitory of spot expansion was 62.35%. Brown spot disease incidence was 88.33% on control and 60.80% on leaves treated by VOCs, lesions expanded slowly and inhibitory rate was 65.75%. SPME GC-MS analysis showed that seven components of the volatiles were identified, all of which are sesquiterpenes with C15H24 structure. They are dihydrocurcumene (CAS NO. 1461-02-5), (E)-β-famesene (CAS NO. 18794-84-8), γ-curcumene (CAS NO. 451-55-8), α-zingiberene (CAS NO. 495-60-3), π-bisabolene (CAS NO. 495-61-4), β-sesquiphellandrene (CAS NO. 20307-83-9) and γ-E-bisabolene (CAS NO. 53585-13-0). When AR03 was cultured for 1 d, the relative content of β-sesquiphellandrene was the highest (80.64%), followed by (E)-β-famesene and α-zingiberene, the relative content was 7.20% and 6.67%, respectively. With the extension of culturing time, the species of each component were the same, but the relative content was different. Except for dihydrocurcumene, the content of other components showed a decreasing trend, when cultured for 6 d, other ingredients decreased more than 50%, besides dihydrocurcumene keeping relatively stable. 【Conclusion】 VOCs produced by B. pumilus AR03 could develop an additive antifungal effect against fungal pathogens on tobacco. B. pumilus AR03 has potential as an important microbial resource for developing antifungal metabolites and new drugs.

    SOIL & FERTILIZER·WATER-SAVING IRRIGATION·AGROECOLOGY & ENVIRONMENT
    Effects of Biochar and Biochar-Based Fertilizer on Soil Microbial Community Structure
    Kun CHEN, XiaoNan XU, Jing PENG, XiaoJie FENG, YaPeng LI, XiuMei ZHAN, XiaoRi HAN
    Scientia Agricultura Sinica. 2018, 51(10):  1920-1930.  doi:10.3864/j.issn.0578-1752.2018.10.011
    Abstract ( 532 )   HTML ( 24 )   PDF (439KB) ( 476 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 Microbes play the key role of switcher in soil nutrient cycling, the aim of this paper was to study the effects of soil microbial community structure by long-term fertilization of biochar and biochar-based fertilizer, and to provide theory reference on reasonable administration of different organic fertilizers, contrasting with traditional organic fertilizer (corn straw and piggery manure compost) in the meantime. 【Method】 This research was based on the long-term experiment of soil improvement of brown soil in Shenyang Agricultural University (began in 2009). The changes of soil physic-chemical properties, soil microbial community structure and their relationship by long-term organic fertilizer plus NPK fertilization were studied by PLFA and correlatively analysis. Soil samples were collected from treatments as farmland with biochar-based compound fertilizer (BF) alone and farmland with piggery manure compost (PMC), corn straw-returning (CS), biochar (BIO) combination of NPK fertilizer, respectively. 【Result】 Soil pH of PMC and BF were higher than BIO significantly; total N (TN) of PMC was significantly higher than BF and CS, which of BIO had no significant difference with PMC; soil organic matter (SOM) of PMC was significantly higher than that of BF and BIO; moisture content (MC) of PMC was the highest of all treatments; total K (TK) of all treatments had no significant difference. Total PLFAs of PMC was significantly higher than other treatments, but there were no significant differences among others; bacteria PLFAs of PMC was the highest of all treatments, which of BF was significantly lower than BIO and CS; fungi, gram-positive, gram-negative PLFAs of PMC were significantly higher than BIO, which of BF had no significant difference with PMC; actinomyces PLFAs of PMC was higher than CS significantly, there were no significant differences between BIO and BF. The result of analysis showed that Shannon-Winner richness index (H) and the fungi/bacteria PLFAs ratio of BF was higher than BIO significantly, the gram- positive/gram-negative PLFAs ratio of BF and PMC were lower than BIO significantly. The result of redundancy analysis (RDA) showed that microbial PLFA was significantly influenced (P<0.01) by soil pH, TN and SOM, and significantly influenced (P<0.05) by MC and TK.【Conclusion】 Soil physico-chemical properties were obvious improved by long-term fertilization of biochar and biochar-based compound fertilizer. Compared to PMC, farmland with BIO was bad for the growth of fungi and gram-negative microbe, and farmland with BIO and BF had different effects on soil microbial community structure, namely, farmland with BIO could increase the biomass of bacteria, while farmland with BF could increase the fungi/bacteria ratio and the diversity of soil microbial community structure. Soil pH, TN, SOM, MC and TK were the important factors which influence the soil microbe community structure in this study in the order.

    Effects of Long-Term Different Fertilization on Sugarcane Yield Stability, Fertilizer Contribution Rate and Nutrition Loss
    HuiPing OU, LiuQiang ZHOU, JinSheng HUANG, Yan ZENG, XiaoHui ZHU, RuLin XIE, HongWei TAN, BiYan HUANG
    Scientia Agricultura Sinica. 2018, 51(10):  1931-1939.  doi:10.3864/j.issn.0578-1752.2018.10.012
    Abstract ( 457 )   HTML ( 5 )   PDF (413KB) ( 512 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 This study was conducted to explore the response of the sugarcane yield stability, fertilizer contribution rate and N, P loss in runoff under long-term different fertilization, with an aim to provide scientific references for establishing the optimal fertilization pattern and promoting the sustainable production of sugarcane as well as the quality of agroecosystem improvement.【Method】 Four different fertilization treatments (non-fertilization (CK), optimum fertilization (OPT), application of 50% N increase based on OPT (OPT+N) and application of 50% P2O5 increase based on OPT (OPT+P)) were chosen from an 8-years fertilization filed experiment. The changes of annual sugarcane stem yield, fertilizer contribution rate and N, P loss in runoff were investigated. 【Result】 At the first 4 years of planting, sugarcane yield declined sharply, and then kept equilibrium around 50 t·hm-2, while fertilization treatments showed a fluctuation in different years, and kept the same trend in the same year. Fertilization significantly increased the sugarcane yield and its stability. With an average of 8 years, sugarcane yield under fertilizer treatments was 70% higher than that under CK. However, there was no significant increase between OPT and OPT+N treatments in sugarcane yield, while OPT was higher than OPT+N treatment. There was no significant difference among fertilizer treatments in sugarcane yield stability. The soil contribution rate in sugarcane field declined sharply at the first 4 years of planting, while fertilizer contribution rate showed the opposite trend, and then both of them basically stabled at around 50%. Both of fertilizer contribution rate and agronomic efficiency in OPT treatment were significantly higher than or equal to OPT+N and OPT+P treatments. Fertilization significantly increased N and P runoff loss. Excessive application of N and P fertilizer significantly increased the corresponding N and P loss, but not N (P) fertilizer loss rate. 【Conclusion】 Excessive application of N and P not only had no advantage in sugarcane yield and its stability, but also resulted in waste of resources and increase of nutrient loss. OPT treatment was a better fertilization model for high and stable yield of sugarcane, high fertilizer contribution rate and low nutrient loss.

    Sources of Cadmium Accumulated in Rice Grain
    Hua YU, YuXian SHANGGUAN, ShiHua TU, YuSheng QIN, Kun CHEN, DaoQuan CHEN, QianCong LIU
    Scientia Agricultura Sinica. 2018, 51(10):  1940-1947.  doi:10.3864/j.issn.0578-1752.2018.10.013
    Abstract ( 363 )   HTML ( 11 )   PDF (350KB) ( 206 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 The objective of this study aimed to understand the sources of cadmium accumulated in the rice grain and to offer scientific support and best timing of agronomic measures to secure safety rice production. 【Method】 The study involved a pot hydroponic experiment with three Cd concentrations of 0, 0.2 and 0.5 mg·L-1 by transplanting uniform rice plants at fully heading stage from a field with Cd contamination. Besides, the rice in the same field were also observed. The rice samples from hydroponic culture were harvested at maturity and those from the field were sampled at full heading stage, milk stage, dough stage and maturity. Dry weight and tissue Cd contents were analyzed for all the samples. 【Result】 The results showed that Cd accumulated in the rice grain was mainly transferred from the root and culm where Cd was stored before full heading in the Cd 0 treatment of the hydroponic culture. When available Cd was low in the soil (i.e., the treatment with rice kept growing in the paddy field), Cd accumulated in the rice grain was from both plant tissues stored Cd before heading and soil Cd uptake by rice after heading. The leaf and chaff appeared as net sources for Cd transferred to the rice grain, while the culm and root behaved as both pools for Cd accumulation and sources to the grain. When the rice grew in relatively high Cd content media (i.e., Cd 0.2 and Cd 0.5 treatments), Cd accumulated in the rice grain was mainly from direct uptake of Cd by the plant and small portions from different rice organs. 【Conclusion】 The Cd accumulated in the grain is transferred from different rice organs before heading stage and from direct uptake from soil/medium; the higher the available Cd in soil/medium, the higher Cd accumulated in the grain; only when there is no available Cd in the soil/medium, the Cd stored in different rice organs becomes the sole source of grain Cd. The culm and root are the two major locations for Cd storage and output in the rice plant. Heading to maturity stages are crucial to control Cd accumulation in rice grain and thus, are key period to secure production of safety rice if proper measures are taken to reduce available Cd in the growth medium as well as its uptake by direct root uptake and transport to the grain.

    HORTICULTURE
    Identification and Expression Analysis Under Abiotic Stresses of OFP Gene Family in Apple
    RuiRui XU, Rui LI, XiaoFei WANG, YuJin HAO
    Scientia Agricultura Sinica. 2018, 51(10):  1948-1959.  doi:10.3864/j.issn.0578-1752.2018.10.014
    Abstract ( 350 )   HTML ( 14 )   PDF (5285KB) ( 336 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 Identification of the OFP (OVATE family protein) genes from apple genome and analysis of gene characteristic, tissue expression pattern and response to abiotic stresses of OFP family genes in apple will be useful to the functional analysis of plant OFP genes. 【Method】 Based on apple genome database, the OFP gene family members were identified and the genes were analyzed using bioinformatics methods. A phylogenetic tree was created using the MEGA5.0 program. Gene structure and chromosomes location were carried out by MapDraw and GSDS separately. Expression pattern analysis of OFP genes in different tissues was done based on the existing microarray database and qRT-PCR. The expression of 13 MdOFPs genes was also analyzed under various stress conditions using qRT-PCR. 【Result】 A total of 28 OFP genes was systematically identified from apple genome and classified into 4 groups including 13, 6, 4 and 5 members according to the gene structure and conserved domain phylogeny relationship. All OFP genes are distributed on 13 apple chromosomes with the largest number six MdOFP on Chr12, suggesting that they have an extensive distribution on the apple chromosomes. Most of the OFP genes have distinctive expression patterns in tissues and response to NaCl and PEG treatment stresses in root and shoot, respectively. MdOFP04 and MdOFP20 were up-regulated obviously in root and shoot, while MdOFP01, MdOFP12 and MdOFP18 have opposite expression pattern in root and shoot under NaCl stress. Temperature stresses significantly regulate the expression of MdOFP and the expression of MdOFP04 and MdOFP17 were significantly increased after high temperature and low temperature stresses. 【Conclusion】 Twenty-eight OFP genes in apple were identified by genome-wide screening. They are classified into four groups and distributed on 13 chromosomes with different tissue patterns and different stress response patterns. These results will be helpful to the functional analysis of OFP genes in apple.

    Morphological Identification and Cultivation of New Germplasm of Apple Homozygous Genotypes
    JiangPeng SHI, ChunFen ZHANG, Shu DENG, LiYuan HOU, Rong XIAO, FuRong LI, YanHui DONG, YuanJun NIE, YiXue WANG, QiuFen CAO
    Scientia Agricultura Sinica. 2018, 51(10):  1960-1971.  doi:10.3864/j.issn.0578-1752.2018.10.015
    Abstract ( 268 )   HTML ( 5 )   PDF (3048KB) ( 133 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 Homozygous genotype lines were obtained by anther culture of different apple pollen grains, so their botanical characteristics, biological characteristics, and their induced culture, root conditions were different. In this study, the ploidy, characteristics and root conditions of each line were observed and analyzed. Appropriate rooting conditions and good combinations of lines for selecting apple homozygous genotypes are of great significance to the innovation of apple germplasm resources.【Method】 The homozygous genotype lines of ‘Gala’, ‘Fuji’ and ‘Red Star’ were obtained by apple anther culture. The ploidy level of homozygous genotype lines was studied using flow cytometry, root and leaf morphology and root conditions of each line were observed and recorded. 【Result】 A total of 32 homozygous lines were obtained by apple anther culture. The results of flow cytometry showed that there were 1 haploid, 1 triploids, 3 tetraploids and 27 diploids. The ploidy rate of 'Red Star' homozygous genotype was the highest, about 28.57%. The root rate, root length and root number of homozygous lines were affected by IBA concentration of rooting medium. When the concentration of IBA was 2-3 mg·L-1, the root features of apple homozygous lines were the best. The leaf number, root length, root number, plant height, leaf length/width and petiole length of each line were different. ‘Red Star’ homozygous genotype lines (DH2-1, DH0-3, DH0-4, DH0-7), ‘Fuji’ homozygous genotype line (DH1-3) and ‘Gala’ homozygous genotype lines (DH2-2, DH2-4, DH2- 12, DH2-20) have a good traits (plants are taller, roots are longer, and the number of roots and leaves are more). The survival rate of ‘Red Star’ homozygous genotype was the highest, about 28.57%. 【Conclusion】 The main ploidy of apple homozygous genotype was diploid. Compared with the diploid lines, the haploid lines had a narrow leaf base, finer petiole, the leaf color and the leaf margin were shallow, while the polyploid lines showed wider leaf base, thicker petioles, leaf color and leaf edge serrated deep.

    Effects of Different Rootstocks on ‘87-1’ Grape Photosynthetic and Chlorophyll Fluorescence Characteristics
    Xiao HAN, HaiBo WANG, XiaoDi WANG, XiaoHao JI, XiangBin SHI, BaoLiang WANG, XiaoCui ZHENG, ZhiQiang WANG, FengZhi LIU
    Scientia Agricultura Sinica. 2018, 51(10):  1972-1981.  doi:10.3864/j.issn.0578-1752.2018.10.016
    Abstract ( 317 )   HTML ( 5 )   PDF (441KB) ( 474 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 The effects of different rootstock on ‘87-1’ grape photosynthetic characteristics and chlorophyll fluorescence characteristics were evaluated, and the results will provide theoretical basis for screening suitable cultivation rootstock.【Method】 Two- and three-year-old ‘87-1’ grape were grafted on eight kinds of rootstocks, including Beta, 1103P, 3309C, 140Ru, 5C, SO4, HuaPu NO.1 and KangZhen NO.1 in 2016 and 2017, and the eight kinds of rootstook-scion combinations were planted in grape center technology demonstration areas of Fruit Research Institute of Chinese Academy of Agricultural Sciences (Xingcheng, Liaoning province, east longitude 120.51° and northern latitude 40.45°). Their spacing in the rows and spacing between rows are 2.5 m×4.0 m. Double plant colonization, oblique dry horizontal dragon shape with horizontal curtain werer applied with the integration of water and fertilizer, and other management measures were routine. The Li-6400 Photosynthetic apparatus was used to measure eight rootstock-scions combinations’ light response curve, carbon dioxide response curve, temperature response curve in the cloudless morning. Before the measurement of three kinds of light curves, the optimal function leaf of different rootstock-scion combinations were selected by Li-6400 Photosynthetic apparatus. Li-6400 Photosynthesis apparatus was also used to choose the leaves with the largest net photosynthetic rate of different rootstock-scion combinations. First of all, the light response curve was measured as the carbon dioxide concentration was set at 400 μmol·mol-1, temperature was set as 25℃, gas velocity was set at 500 mmol·s-1, photosynthetically active radiation was set at 2 000, 1 800, 1 500, 1 200, 800, 400, 200, 100, 50, 20, 0 μmol·m-2·s-1 from strong to weak. Secondly, the carbon dioxide response curve was measured as the photosynthetically active radiation was set at 1 200 μmol·m-2·s-1, temperature was set at 25℃, gas velocity was set at 500 mmol?s-1, carbon dioxide concentrations were set at 2 000, 1 800, 1 500, 1 200, 800, 400, 200, 150, 100, 50, 20 μmol·mol-1 from high to low. Finally, the net photosynthetic rate curve was measured as the photosynthetically active radiation was set at 1 200 μmol·m-2·s-1, carbon dioxide concentration was set at 400 μmol·mol-1, gas velocity was set at 500 mmol·s-1, the temperatures were set at 25, 27, 30, 32, 35,27℃ from low to high. And then, the apparent quantum yield, light compensation point, carboxylation efficiency, carbon dioxide compensation point, dark respiration rate were obtained using right angle hyperbolic correction model. And the FMS-2 pulse-modulated fluorometer was used to obtain minimal fluorescence, maximal fluorescence, variable fluorescence, potential quantum yield of PSⅡ, maximal photochemical efficiency of PSⅡ in the dark. Then, variance analysis was used to compare the effects of different stock on photosynthetic parameters of ‘87-1’ grape, and Topsis comprehensive evaluation method was used to rank the eight rootstock-scion. 【Result】 The results of Topsis comprehensive evaluation showed that rootstock-scion combinations of ‘87-1’/3309C and ‘87-1’/1103P were ranked the first and second to tolerate weak light,and with the low light compensation point, high apparent quantum yield, low dark respiration rate. The rootstock-scion combinations of ‘87-1’/3309C and ‘87-1’/Huapu NO.1 were ranked the first and second to tolerate low carbon dioxide levels,and with low carbon dioxide compensation point, high apparent quantum yield. The rootstock-scion combinations of ‘87-1’/SO4, ‘87-1’/Huapu NO.1 were ranked the first and second to tolerate high temperature,and with little net photosynthetic changes at different temperatures and high net photosynthetic rate at high temperature. Furthermore, the results of variance analysis showed that the highest minimal fluorescence was found in ‘87-1’/140Ru and ‘87-1’/SO4 rootstock-scion combinations, the highest of variable fluorescence was found in ‘87-1’/1103P rootstock-scion combination, the highest of rate variable fluorescence and minimal fluorescence were found in ‘87-1’/1103P and ‘87-1’/3309C two kinds of rootstock-scion combinations. There was no significant difference of maximal fluorescence,the rate of variable fluorescence and maximal fluorescence among all 8 kinds of rootsock-scion combinations had no. 【Conclusion】 The rootstocks 3309C and 1103P can improve ‘87-1’ grape weak light tolerance ability efficiently, the rootstock 3309C and Huapu NO.1 can improve ‘87-1’ grape low carbon dioxide level tolerance ability efficiently, the rootstocks SO4 and Huapu NO.1 can improve ‘87-1’ grape high temperature tolerance ability efficiently, the rootstocks 1103P and 3309C can improve the beneficial to the enhancement of ‘87-1’ grape the primary light energy conversion efficiency.

    FOOD SCIENCE AND ENGINEERING
    The Solid-State Fermentation Process of Rhodotorula Mucilaginosa for Producing Carotenoids
    JiaJing SUN, Mao LI, ZhiHong SUN, ZhiRu TANG, XiangXin ZHANG, JinChao CHEN
    Scientia Agricultura Sinica. 2018, 51(10):  1982-1994.  doi:10.3864/j.issn.0578-1752.2018.10.017
    Abstract ( 349 )   HTML ( 8 )   PDF (434KB) ( 477 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 This study was performed to enhance carotenoid yield, to improve nutritional value of fermentation product, and to reduce the production cost of carotenoids through optimizing solid-state fermentation substrate and fermentation conditions of Rhodotorula mucilaginosa.【Method】 In this study, Rhodotorula mucilaginosa TZR2014 was used as a inoculant. First, the Mixture-Design of Design-Expert software was used to design the fermentation substrate, and the contents of ingredients as followed: 50%-80% wheat bran, 6%-20% soybean meal, 3%-15% maize flour, 2%-14% rice bran, 2%-10% maize syrup, 0.4%-2.5% ammonium sulfate, 0.05%-0.5% monopotassium phosphate, and 0.03%-0.3% magnesium sulfate. Then the optimal ratio of ingredients in substrate was determined according to the carotenoid yield. Based on this result, an L16(45) orthogonal design was used to optimize the fermentation conditions, including inoculum (5.0%-12.5%), fermentation time (60.0-96.0 h), fermentation temperature (26-32℃), and fermentation pH (60.0%-75.0%). Finally, the number of Rhodotorula mucilaginosa and contents of carotenoids, crude fiber, crude protein, water, crude fat, ash, calcium, phosphorus, and amino acids in fermentation product were determined to evaluate the effects of the optimized fermentation process on the nutritional values of fermentation product. 【Result】 The results showed that there was a positive correlation between maize starch content in substrate and carotenoid content in fermentation product (r=0.344, P=0.040) or between rice bran content in fermentation substrate and carotenoid content in fermentation product (r=0.329, P=0.050). There was a significantly negative correlation between carotenoid yield and the content of wheat bran in solid-state fermentation substrate (r=-0.336, P=0.045). There was a positive correlation between the number of live bacteria of Rhodotorula mucilaginosa in fermentation product and the content of soybean meal in fermentation substrate (r=0.510, P=0.001). Inoculum, fermentation temperature, pH, and moisture had extremely significant impacts on the number of Rhodotorula mucilaginosa (P<0.01), thereinto, fermentation temperature had the greatest effect on the number of Rhodotorula mucilaginosa, followed by moisture, inoculum, and pH. Fermentation time, fermentation temperature, and pH had extremely significant influences on the carotenoid content in the fermentention product (P<0.01), and fermentation temperature had the greatest influence on the carotenoid content in the fermented product, followed by pH and fermentation time. After the optimization of the fermentation process, the carotenoid yield by Rhodotorula mucilaginosa TZR2014 was increased to 4 535 μg·kg-1; the bacteria number was increased to 3.79×109 CFU/kg; the contents of crude fiber, crude protein, ash, threonine, glutamate, and proline in fermentation product were significantly increased (P<0.05), meanwhile, the contents of histidine, water, and crude fat was significantly decreased (P<0.05). 【Conclusion】 The optimal ratio of solid-state fermentation substrate for Rhodotorula mucilaginosa was as followed: 52.5% wheat bran, 20.0% soybean meal, 3.0% maize flour, 14.0% rice bran, 10.0% maize syrup, 0.4% ammonium sulfate, 0.05% monopotassium phosphate, and 0.04% magnesium sulfate. The optimal fermentation conditions were as followed: inoculum 5.0%, fermentation time 72 h, fermentation temperature 28.0℃, pH 6.0, and moisture 60.0%. The results suggested that the optimized fermentation process of Rhodotorula mucilaginosa enhanced the yield of carotenoids and improved the nutritional value of fermentation product.

    ANIMAL SCIENCE·VETERINARY SCIENCE
    The E2 Protein Mediates the Attachment and Internalization of Classical Swine Fever Virus in Cells
    CaiXia YIN, ShaoXiong YU, Kun SONG, Su LI, YuYing YANG, HuaJi QIU
    Scientia Agricultura Sinica. 2018, 51(10):  1995-2003.  doi:10.3864/j.issn.0578-1752.2018.10.018
    Abstract ( 482 )   HTML ( 8 )   PDF (1409KB) ( 165 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 It has been shown that the entry of classical swine fever virus (CSFV) into host cells is mediated by clathrin-mediated endocytosis. However, the viral protein involved in entry stage remains to be elucidated. This study aimed to clarify the role of the E2 protein in the entry of CSFV. 【Method】 Lentivirus carrying the E2 gene were packaged by transient transfection of HEK-293T cells. A cell line stably expressing the E2 protein was established through transducing the suspension 293 cell with the lentiviruses. The expression of the recombinant E2 (rE2) protein was optimized to achieve higher production, and the rE2 protein was purified by affinity chromatography. The expression level and reactivity of the rE2 protein was identified by SDS-PAGE and Western blotting. The effects of the E2 protein on CSFV infection, attachment and internalization were determined by respective assays. Polyclonal anti-E2 antibodies were prepared by immunizing BALB/c mice with the rE2 protein, and its blocking rate was determined by blocking ELISA. Attachment and internalization assays were performed using the prepared polyclonal antibodies to demonstrate the role of the E2 protein in virus attachment and internalization. 【Result】 The cell line 293 was successfully transduced with the lentivirus vector. The SDS-PAGE produced the anticipated band size of rE2 no matter the sample was treated with the reducing agent or not. Western blotting results showed that the rE2 protein could be recognized by the anti-E2 monoclonal antibody WH303. The results indicated that suspension 293 cell line could stably express the rE2 protein. Under the optimized expression condition, the concentration of the rE2 protein in the supernatants of the established suspension 293 cell line was up to 5.84 μg·mL-1. Soluble protein blocking assay results showed that the expressed rE2 protein exerted antiviral activity during the process of CSFV infection. CSFV infection were significantly inhibited by the rE2 protein treated in the entry step; Blocking ELISA and serum neutralization test showed that the anti-E2 polyclonal antibodies could neutralize CSFV infection; at the same time, attachment and internalization assays demonstrated that CSFV internalization could be inhibited by the rE2 protein and viral attachment was blocked by the polyclonal antibodies. 【Conclusion】 The E2 protein was involved in attachment and internalization of CSFV.

    Study on Production Performance of Small-Tail Han Sheep and Different Combinations of Hybrid Sheep Fed Whole-Plant Corn Silage
    Jie ZHANG, Chen ZHANG, ChongYu ZHANG, XuXian QU, WangTao ZHAN, JingChao YANG, YingNan WANG, GuiGuo ZHANG
    Scientia Agricultura Sinica. 2018, 51(10):  2004-2012.  doi:10.3864/j.issn.0578-1752.2018.10.019
    Abstract ( 333 )   HTML ( 16 )   PDF (385KB) ( 204 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 The aim of this study was to explore the effects of the total mixed ration (TMR) based on the whole-plant corn silage on the production performance and economic profit of fattening sheep of different varieties and therefore offer theoretical support to the application of the whole-plant corn silage and healthy sheep production. 【Method】 Nutritional composition and rumen degradation characteristics of the whole-plant corn before and after ensiling were evaluated by in Sacco method. Four varieties of 3-month-old healthy male lambs (breast-milk weaned) with similar body weight (27.6±1.7) kg, were selected. There were the following four treatments accordingly: Small-Tail Han (SH), “Dorper×Small-Tail Han” F1 (DH), “Mongolia×Han” F1 (MH) and “Dorper×Wadi” F1 (DW). Each treatment had 30 lambs, which were divided into 3 replications. Forage to concentrate ratio of TMR is 25:75. There was an adaptation period of 10 days followed by an experimental period of 80 days, measuring the growth performance and slaughtering performance. 【Result】 Comparing to the whole-plant corn before ensiling, the silage did not have different content of crude protein (CP) (P>0.05) but owned greater DM, OM and rapidly degraded fraction (a) (P<0.01) and lower NDF, ADF and effective degradation rate (ED) (P<0.001). There was a tendency of higher DMI with hybrid sheep than with SH (0.05<P<0.1). The average daily gain (ADG), feed conversion ratio (FCR), carcass weight, neat weight, dressing percentage, neat percentage and neat percentage expressed in relation to carcass weight of DH and DW sheep were significantly higher than those of SH sheep (P<0.05). There was no significant difference in feed cost among all groups (P>0.05). The net meat gain, meat production efficiency and economic profit of DH and DW sheep were significantly higher than those of SH and MH sheep (P<0.01). 【Conclusion】 The whole-plant corn silage was suitable for use as the main feed source of TMR for fattening sheep, and all different breeds of sheep showed an expected production performance. The DH and DW hybrid sheep had a higher production performance and economic profit than that of SH sheep and MH hybrid sheep. However, there was no difference in production between SH and MH crossbred sheep. Therefore, DH and DW hybrid sheep were more suitable as fattening breeds in Huang-Huai-Hai farming area.

    RESEARCH NOTES
    Development of a Real-Time Fluorescent Quantitative PCR Method for the Detection of Tomato chlorosis virus and Its Application
    TianBo DING, XiaoBei LIU, Jie LI, KeKe WEI, Dong CHU
    Scientia Agricultura Sinica. 2018, 51(10):  2013-2022.  doi:10.3864/j.issn.0578-1752.2018.10.020
    Abstract ( 360 )   HTML ( 7 )   PDF (1642KB) ( 584 )   Save
    Figures and Tables | References | Related Articles | Metrics

    【Objective】 The objective of this study is to develop a real-time fluorescent quantitative PCR (RT-qPCR) method to detect the Tomato chlorosis virus (ToCV) and its concentration in plant and the vector Bemesia tabaci.【Method】 Firstly, the special primers ToCVqF/R were designed using Primer 3.0 for RT-qPCR based on the highly conserved region of the minor coat protein (CPm). The primers specificity was evaluated by Primer-BLAST. Secondly, the fragment was amplified using the primers ToCVqF/R by PCR within the ToCV-infected, Tomato yellow leaf curl virus (TYLCV)-infected, and Tomato spotted wilt virus (TSWV)-infected tomato samples, and cloned into pMDTM 18-T Vector. The positive clone was selected and multiplied in LB liquid medium containing ampicillin. Furthermore, the recombinant plasmid was extracted from the clone and served as the ToCV standard. Then, the standard curve was generated for quantitative analysis using the ten-fold dilution of the recombinant plasmid. In addition, the detection sensitivity of RT-qPCR was also evaluated and compared to that of reverse transcription-polymerase chain reaction (RT-PCR). Finally, this method was used to detect and quantify ToCV in eight tomato samples with suspected ToCV infection and single B. tabaci after acquiring ToCV for 24 h, according to the amplification curves, melting curves and Ct values.【Result】 The purpose strips could be successfully amplified with the primers ToCVqF/R only in the ToCV-infected tomato sample and positive control, which indicated that the primers was highly specific. Additionally, the sequencing result of the fragment was in according with the target gene CPm. The ToCV standard curve showed that the Ct values gradually increased as the concentration of recombinant plasmid decreased. Among the concentration range of 2.7×103-2.7×109 copies/μL, the standard curve showed a good linear relationship between Ct values and plasmid concentration. The correlation coefficient of the standard curve was 0.9911, and the amplification efficiency was 100%. The Straight-line equation is y=-3.32×lgx+40.06 (y and x represent the Ct value and plasmid concentration, respectively). The sensitivity of RT-qPCR was 100 times higher than regular RT-PCR. The minimum detectable concentration of ToCV plasmid standard in RT-qPCR assay is 2.7×103 copies/μL, while that in RT-PCR assay is 2.7×105 copies/μL. This method could successfully detect the ToCV within tomato samples, and the result was consistent with RT-PCR assay. Six of eight tomato samples were infected by ToCV, the viral titers of which were 2.48×105, 2.21×105, 7.97×104, 3.74×107, 3.37×107 and 2.78×106 copies/μL, respectively. After 24 h feeding on ToCV infected tomato plants, 100% of the B. tabaci acquired this virus successfully, and the virus concentration in single whitefly ranged from 6.46×102 to 2.55×104 copies/μL.【Conclusion】 The RT-qPCR method is applicable for accurate and sensitive detection of ToCV both in plants and vector insects, which will provide a technical support for the monitoring and early warning of this virus.