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Table of Content

    01 October 2015, Volume 48 Issue 19
    CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS
    Molecular Detection for Quality Traits-Related Genes in Yangmai Series Wheat Cultivars
    ZHANG Xiao, ZHANG Bo-qiao, JIANG Wei, Lü Guo-feng, ZHANG Xiao-xiang, LI Man, GAO De-rong
    Scientia Agricultura Sinica. 2015, 48(19):  3779-3793.  doi:10.3864/j.issn.0578-1752.2015.19.001
    Abstract ( 454 )   HTML ( 7 )   PDF (1099KB) ( 510 )   Save
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    【Objective】The purpose of this study is to clarify the distribution of quality-related genes in Yangmai series wheat varieties and provide guidance for wheat genetic breeding and production. 【Method】Single pedigree of twenty-one Yangmai wheat varieties were selected. Grain hardness was measured by single grain analyzer SKCS-4100. Hardness gene, low molecular weight glutenin subunit gene (LMW-GS), Wx gene coding key enzyme for amylose synthesis, polyphenol oxidase (PPO) gene, yellow pigment content (PSY) gene and pre-harvest sprouting tolerance (Vp1) gene were detected in twenty-one Yangmai wheat varieties by their corresponding functional markers through polyacrylamide gel electrophoresis technology, while high molecular weight glutenin subunits(HMW-GS)and Wx protein subunits composition were detected by SDS-PAGE. 【Result】There are sixteen soft wheat through grain hardness analyzer, which accounted for 76.19%, however, the hard and mixed wheat only accounted for 19.05% and 4.76%, respectively. In hardness and mixed wheat, there are four varieties with pinb-D1b mutation, but in sixteen soft wheat, no mutation of Pinb-D1b was found. When detecting the composition of HMW-GS, allele 1 and Null accounted for 38.10% and 61.90%, respectively, at the Glu-A1 locus, and allele 7+8 and 7+9 at Glu-B1 took up 57.14% and 42.86%, respectively, whereas at the Glu-D1 locus, allele 2+12 was 85.71%, 5+10 was 14.29%, respectively. Meanwhile, for LMW-GS compositions, “Glu-A3c, Glu-B3g” was the main genotype. Genotype frequencies of Glu-A3c and Glu-A3d were 90.48% and 9.52%, respectively. Glu-B3g and Glu-B3i frequencies were 95.24% and 4.76%, respectively. Molecular detection indicated that the allele variation Wx-B1b which resulted inthe subunit Wx-B1 deletion was only found in Yangmai13. However, SDS-PAGE method turned out that Yangmai13 and Yangmai5 both lacked Wx-B1 subunit. Results also showed that, in all twenty-one Yangmai varieties, at 2AL locus for PPO activity, the frequency of Ppo-A1a for high PPO activity was 52.38%, and Ppo-A1b related low PPO activity was 42.86%. Apart from Ppo-A1b, another low PPO related gene Ppo-D1 located on 2DL, whose frequency was 90.48%. It was also found that phytoene synthase genes Psy-A1a and Psy-A1b accounted for 19.05% and 80.95%, respectively. When detecting pre-harvest sprouting tolerance gene with marker Vp1B3, tolerant genotype Vp1Bc and susceptible genotype Vp1Bawere amplified in this report, accounting for 90.48% and 9.52%, respectively.【Conclusion】Most Yangmai varieties were soft wheat, which was due to the pinb-D1a genotype at pinb-D1 locus, Null and 2 + 12 at Glu-A1 and Glu-D1 loci and Glu-A3c at Glu-A3 locus. These varieties can be applied as donor parents in weak gluten wheat breeding. Quality of middle-gluten wheat Yangmai15 and Yangmai16 were superior mostly because of the Pinb-D1 locus mutation, so the selection of Pinb-D1 mutation should be enhanced in quality improvement breeding for middle-gluten wheat. Yangmai1, Yangmai4, Yangmai5, Yangmai9, Yangmai18, Yangmai19 and Yangmai22 with low polyphenol oxidase activities and low yellow pigment content could be used as donors for selecting materials with high flour whiteness and good color. Yangmai158, Yangmai10, Yangmai16 and Yangmai17 varieties with Pinb-D1b allele, Yangmai2, Yangmai4 and Yangmai5 with subunit pair of “5+10” at Glu-D1 locus, Yangmai13 and Yangmai5 with Wx-B1 deletion, could be applied to the protein and starch quality improvement of middle-gluten wheat for their high quality traits.
    Cloning and Functional Analysis of Cotton U6 Promoter with High Transcription Activity in Cotton Pollen
    LEI Jian-feng, WU Juan, CHEN Xiao-jun, YU Tian-ping, NI Zhi-yong, LI Yue, ZHANG Ju-song, LIU Xiao-dong(College of Agronomy, Xinjiang Agricultural University/Research Center of Cotton Engineering, Mini
    Scientia Agricultura Sinica. 2015, 48(19):  3794-3802.  doi:10.3864/j.issn.0578-1752.2015.19.002
    Abstract ( 700 )   HTML ( 6 )   PDF (1959KB) ( 1392 )   Save
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    【Objective】In order to take advantage of the genome editing technology in cotton molecular breeding, U6 promoters were cloned from cotton variety Xinhai16 (Gossypium barbadense L.) and then a GbU6 promoter was selected with high transcription activity in cotton germ cells (pollen), which will lay an important foundation for cotton molecular breeding.【Method】Two rounds of PCR were adopted to clone accurately the full-length of GbU6 promoter without redundant sequence at 3’ end, five pairs of primers were designed using software Primer Premier 5.0. The first round PCR was carried out to obtain DNA fragments which cover five full-length GbU6 promoters, respectively. After confirming by sequencing, five GbU6 promoters were subcloned precisely into CRISPR/Cas9 genome editing vector by the second round of PCR (transfer PCR). After right sequencing of the transfer PCR products, the necessary element for transcriptional function of 5 kinds of GbU6 promoter was analyzed using DNAMAN software. Then GUS report gene was cloned using pBI101 plasmid as template. After confirming by sequencing, GUS gene was cloned into CRISPR/Cas9 genome editing vectors carrying five GbU6 promoters, respectively, by BbsⅠdigestion and ligation, which create five kinds of fusion expression vectors of GbU6::GUS. DNA fragments of CaMV35S promoter-driven GUS and fiveGbU6::GUS were obtained through High fidelity PCR and then were transferred into cotton pollen using a particle gun. Results were observed using a stereomicroscope after GUS staining. Each transformation was repeated three times. 【Result】Five different GbU6 promoters were cloned after two rounds of PCR and they contain 1 166 bp, 1 119 bp, 1 134 bp, 1 214 bp and 1 176 bp, respectively. Construction of the corresponding five CRISPR/Cas9 genome editing vector with five different GbU6 promoters were done. After sequence comparation of U6 promoter between Arabidopsis and cotton, results showed that cotton U6 promoter contained the conserved -60 USE motif and -30 TATA box and the distance between two elements also was fixed, just like Arabidopsis U6 promoter. The results of transient transformation showed that four cloned GbU6 promoters could drive the expression of GUS gene in cotton pollen which were stained into blue. Among the four promoters, the GbU6-5P::GUS exhibited deeper blue, similar to the CaMV35S promoter.【Conclusion】GbU6 promoters with high-level transcription in cotton pollen were cloned, thus providing an efficient promoter for genome editing technology of CRISPR/Cas9 in cotton.
    TILLAGE & CULTIVATION·PHYSIOLOGY & BIOCHEMISTRY·AGRICULTURE INFORMATION TECHNOLOGY
    Effects of Wheat Stubble Height on Growth and Water Use Efficiency of Mechanized Sowing Summer Maize
    GAO Ying-bo, TAO Hong-bin, ZHU Jin-cheng, HUANG Shou-bing, XU Cai-long, SHENG Yao-hui, WANG Pu
    Scientia Agricultura Sinica. 2015, 48(19):  3803-3810.  doi:10.3864/j.issn.0578-1752.2015.19.003
    Abstract ( 544 )   HTML ( 2 )   PDF (408KB) ( 479 )   Save
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    【Objective】In winter wheat-summer maize rotation system, wheat stubble was retained in field after harvest and maize seeds were sowed directly without tillage. Standing stubble height affected light penetration and straw residue amounts, which might affect soil water humidity, maize emergence rate and crop development. This research aimed at finding out a proper standing stubble height to increase water use efficiency and emergence rate, and finally to improve grain yield of summer maize.【Method】Summer maize hybrid Zhengdan958 was used as experimental materials. A 3yr (2012-2014) field experiment was conducted in order to explore a proper stubble height at Wuqiao Experimental Station of China Agricultural University, Hebei Province. Three stubble height treatments were compared: 35, 25, and 15 cm. The emergence rate, crop growth parameters, dry matter accumulation, soil water consumption and water use efficiency of summer maize were measured in this research. 【Result】From sowing to jointing stage, the treatment with stubble height 35 cm significantly reduced soil water consumption amount, while emergence rate and plant height uniformity of summer maize were both increased significantly. Plant height and leaf area of 35 cm stubble were both greater while stems were thinner than under shorter stubble height treatments at 4-leaf stage. At jointing stage, plant height, stem diameter, leaf area and dry biomass of 35 cm stubble height were all the greatest among treatments. Summer maize yield of 35 cm stubble height was 10.59% and 5.31% higher than that of 25 and 15 cm. Increased grain yields were the result of the increase of kernels number per unit area. Moreover, water use efficiency was 21.55% and 12.64% greater than that of 25 and 15 cm, respectively. The water use efficiency showed large differences between annual precipitations. During the years with more rainfall, water use efficiency was low.【Conclusion】Wheat stubble height of 35 cm was helpful to maize seedling growth and strongly improved the grain yield and water use efficiency of summer maize.
    The Physiological Characteristics of the Antioxidant System of Colored Cotton and the Effects of Cellulose Accumulation on Cotton Fiber Quality
    SONG Shi-jia, SUN Hong-chun, ZHANG Yong-jiang, LIU Lian-tao, BAI Zhi-ying, LI Cun-dong
    Scientia Agricultura Sinica. 2015, 48(19):  3811-3820.  doi:10.3864/j.issn.0578-1752.2015.19.004
    Abstract ( 249 )   HTML ( 1 )   PDF (384KB) ( 467 )   Save
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    【Objective】To research the differences of colored cotton fiber quality and variation in physiological activities of boll development, an experiment was conducted to explore the main period and related influencing factors of fiber quality formation in the Yellow River Valley cotton-farming region of China. This study will provide a theoretical basis for quality improvement of colored cotton. 【Method】In field conditions, three colored cotton varieties, Zhongmian 51 (light brown, CCRI51, hybrid cotton), Zhongmian 81 (dark brown, CCRI81), Zhongmian 82 (dark green, CCRI82), and one ordinary white cotton variety Guoxin 3 (GX3) were used as experimental materials. The cotton flowers grown at 1-2 nodes of 5-6 fruit branches were marked in each district on the same day. Then, fiber quality of the marked bolls was tested at harvest time. In addition, the marked bolls were divided into boll shell, cotton seed and fiber in the 10 DPA (days post anthesis), 20 DPA, 30 DPA, 40 DPA, separately. For each organ of the marked bolls, indexes such as soluble protein content, POD activity, SOD activity, MDA content, and cellulose content of cotton fiber were measured. And fiber qualities, bolls’ development characteristics of different varieties, and the correlation between both were studied.【Result】CCRI 51, as a hybrid cotton variety, had the highest upper fiber length, uniformity, and strength. And the order of that of the other conventional cotton was GX 3>CCRI 81 >CCRI 82. Micronaire value of GX 3 was significantly higher than that of the other cotton varieties. Soluble protein content in boll shell of GX 3 was higher than that in the other varieties in the boll development. Meanwhile, a significant difference was observed among varieties in each period. The largest decreased ratio of fiber soluble protein content of GX 3 appeared in the early time after flowering (10-20 DPA, 66.67%). However, for the other varieties, it appeared at the later stage after flowering (20-40 DPA, 23.08%-61.19%). POD activity of boll shell of all varieties rose in the 10-20 DPA. And that of the variety CCRI 82 was the highest. POD activity in boll shell of GX 3 and CCRI 51 was significantly higher than that of CCRI 81 and CCRI 82. In the 10 DPA, SOD activities in boll shell and cotton seed of GX 3 both were the highest in the 4 varieties. SOD activity in boll shell of CCRI 51 and CCRI 81 showed a gradual increasing trend during its development. SOD activity in cotton seed of CCRI 51 in the 40 DPA increased by 53.18% compared with that in the 30 DPA; also, the value was significantly higher than the other varieties. However, for CCRI 82 fiber, SOD activity declined time occurred 10 days earlier than that of the other varieties. MDA content of GX 3 cotton seed showed a decreasing trend in the 20-30 DPA. However, that of the other varieties showed an increasing trend in this period. The cellulose speedy accumulation time of CCRI 51 occurred earlier and longer than the other varieties. For GX 3, cellulose speedy accumulation time was 3 days earlier than that of CCRI 81 and CCRI 82, and its speedy accumulation duration was 1-5 days longer than that of CCRI 81 and CCRI 82. MDA content in cotton seed in the 10 DPA showed a significantly negative correlation with fiber length and fiber strength, and the coefficients were -0.964 and -0.968, respectively. POD activity in the boll shell in 10 DPA was significantly and positively correlated with fiber length. And cellulose speedy accumulation duration was significantly and positively correlated with fiber length and strength. Protein content in fiber in 20 DPA, MDA content in cotton seed in 30 DPA and cellulose speedy accumulation time were significantly and negatively correlated with fiber uniformity. MDA content in cotton seed in 20 DPA, protein content of boll shell in 30 DPA and MDA content in 40 DPA were all significantly and positively correlated with micronaire value. Both protein content in cotton seed in 40 DPA and POD activity in fiber in 20 DPA were significantly and positively correlated with tensile stretch. 【Conclusion】 Compared to white cotton, colored cotton bolls had a poor antioxidant enzyme system activity, which affected the formation and growth of the primary cell wall of fiber in 0-20 DPA, and promoted coloring substances deposited in the secondary cell wall of fiber, which affected the cellulose accumulation in secondary cell wall of fiber. Shorter cellulose accumulation duration and a late cellulose speedy accumulation time of colored cotton were both the direct cause of low fiber quality.
    Comparison of Root Morphological and Activity of Representative Soybean Cultivars (MG III) Developed in the USA and China
    GUO Xiao-hong, WANG Xing-cai, MENG Tian, ZHANG Hui-jun, AO Xue, WANG Hai-ying, XIE Fu-ti
    Scientia Agricultura Sinica. 2015, 48(19):  3821-3833.  doi:10.3864/j.issn.0578-1752.2015.19.005
    Abstract ( 370 )   HTML ( 5 )   PDF (375KB) ( 455 )   Save
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    【Objective】The research was compared on the root morphological and activity of soybeans [Glycine max (L.) Merr.] developed in different years from the different breeding programs of Ohio, USA, and Liaoning, China, which were at the same latitude. The purpose of this study was to reveal the evolutionary trend of root traits, and the response to the fertilizer rate with the improvement of seed yield. 【Method】 The pot experiment was conducted in 2012 and 2013, and used a completely randomized design with three replications, and all used cultivars had the same ancestors (Williams and Amsoy). The diammonium phosphate was applied at the seedling stage (V2) with a solution condition, and had three levels (0, 150, and 300 mg·kg-1dry soil). The plants were cut at the cotyledonary node at the stages of V4, R2, R6, and R7, and the root bleeding sap weight was measured by a weight difference method with absorbent cotton suction. The plant root was scanned by an Epson Expression 10000XL, and then WinRhizo2012 software analyzed the scanning images to determine root length, root surface area, and number of root hairs, and the root activity was measured by a triphenyl tetrazolium chloride method (TTC). The root samples were dried in the oven at 105?C for 30 minutes and at 80℃ at least for 72 hours to achieve a constant weight, and the seed yield per plant was measured at the mature stage. 【Result】 The root length, root surface area, number of root hairs, root dry weight, root bleeding sap weight, and root activity of Ohio and Liaoning cultivars had the same improvement trend, and all of those were increased with the yield improvement, especially at the stage of R6 and R7, and the value of root morphological and activity was increased greatly by years. Compared with the cultivars of different regions, the root length, the improvement progress of root surface area, the number of root hairs, the root bleeding sap weight, and the root activity of Ohio cultivars was better than the Liaoning cultivars. With the increasing of fertilization rates, the root traits value of the current cultivars of Ohio and Liaoning showed very well at both the medium (150 mg·kg-1dry soil) and high (300 mg·kg-1dry soil) fertilization levels, and those of the middle cultivars of two regions were best at the medium fertilization rate. Under the same fertilizer rate, the root traits value of current cultivars was greater than the ancestor and middle cultivars. The results of correlation and path-coefficient showed that root length, root surface area, root dry weight, root bleeding sap weight, and root activity had significant positive correction with the seed yield per plant at the stages of V4, R2, R6, and R7, and that the correction between root activity and seed yield per plant was the best. 【Conclusion】During the improvement of the soybean yield of the Ohio and Liaoning cultivars, the root morphological and activity had the same improvement trend, and the root length, root surface area, root bleeding sap weight, and root activity were increased with the yield improvement, but the improvement progress of Ohio cultivars was greater than that of Liaoning cultivars. Fertilizer tolerance was enhanced in both Ohio and Liaoning cultivars, the root traits values of current cultivars of two regions showed very well at both the medium (150 mg·kg-1dry soil) and high (300 mg·kg-1dry soil) fertilization levels, and that of the middle cultivars were best at the medium fertilization rate.
    PLANT PROTECTION
    Postulation of Stripe Rust Resistance Genes in Chinese 40 Wheat Landraces and 40 Commercial Cultivars in the Southern Region of Gansu Province
    WANG Tu-hong, GUO Qing-yun, LIN Rui-ming, YAO Qiang, FENG Jing, WANG Feng-tao, CHEN Wan-quan, XU Shi-chang
    Scientia Agricultura Sinica. 2015, 48(19):  3834-3847.  doi:10.3864/j.issn.0578-1752.2015.19.006
    Abstract ( 488 )   HTML ( 2 )   PDF (386KB) ( 367 )   Save
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    【Objective】 The objective of this study is to determine the types of stripe rust resistance, the potential resistance genes and the genetic diversity in Chinese 40 wheat landraces and 40 commercial cultivars grown in Gansu Province, that will provide reference information for the selection and utilization of stripe rust resistance resources. 【Method】 For stripe rust resistance identification of 80 wheat accessions, the prevalent race CYR32 of Puccinia striiformis f. sp. tritici and 12 mixed races including prevalent races and important pathogenic isolates were used for inoculation at the seedling stage in greenhouse and adult-plant stage in nursery field, respectively. Thirty wheat lines with known resistance genes, 17 international and European stripe rust differential varieties, 80 landraces and commercial cultivars and susceptible control Mingxian 169 were inoculated with 25 races of different virulences of P. striiformis at the seedling stage in greenhouse. Besides the comparison of resistance spectrum to stripe rust and pedigree of these wheat cultivars, the tested cultivars and standard cultivars were compared to postulate the possible resistance genes or their combinations contained in the landraces and commercial wheat cultivars according to their infection types to different races. For genetically classifying the cultivars using their pedigree information, the genetic similarity coefficient was calculated by software NTSYSpc-2.2, and a distance tree was constructed using UPGMA (unweighted pair-group mean average). 【Result】 Most of the tested cultivars showed a high level of stripe rust resistance at adult-plant stage, except that Qingnong 1, Qingnong 2 and Hongkexiaomai 2 were susceptible. Twenty cultivars such as Lantian 3, Lantian 4 and Lantian 6 exhibited slow rusting resistance, and the other cultivars ranged in resistance from moderate level to immunity. The resistance phynotype of the tested cultivars was mainly provided by all-stage resistance and partly by adult-plant resistance. Most commercial cultivars from Gansu Province contain Yr9, Yr24 and/or Yr26, and some of them also contain other unknown genes. For instance, Yr9 was postulated to be present in cultivars Lantian 1, Lantian 14, Lantian 17, Lantian 21, Qingnong 4; Yr24 in cultivars Lantian 24, Zhongliang 04335 and Tianxuan 51; and Yr26 in cultivars Lantian 17, Lantian 23, Lantian 25, Zhongliang 17, Zhongliang 04260 and Tianxuan 43, Tianxuan 48. Among the landraces, 19 of them were found to carry unknown resistance gene(s) alone and the other landraces may contain Yr1, Yr2, Yr4, Yr6, Yr7, Yr8 and/or Yr40. Results of the cluster analysis also indicated that the commercial wheat cultivars possessed higher levels of genetic similarity and had wider resistance spectra than the landraces. The similar coefficient of resistance types among the 40 commercial wheat cultivars varied from 0.30 to 1.00. They were classified into 3 groups when the similar coefficient value was 0.70: one group included Lantian 15 alone; the other group included Qingnong 1 and Qingnong 2; and the third one included 37 cultivars. But the range of resistance spectrum was different among the landraces, which possess a higher level of genetic diversity. The similar coefficient of resistance types was from 0.38 to 1.00, and the 40 cultivars were classified into 9 groups when the similar coefficient value was 0.70. 【Conclusion】All tested wheat cultivars showed a higher level of stripe rust resistance. Compared to the commercial wheat cultivars in Gansu, landraces possessed abundant genetic diversity and more effective stripe rust resistance gene(s), so they can be used as stripe rust resistance resources in wheat breeding programs.
    Effects of FBT on Induction of Systemic Resistance in Cucumber Against Cucumber Fusarium Wilt Caused by Fusarium oxysporum f. sp. cucumerinum Owen
    SHI Yan-xia, XU Yu-fang, XIE Xue-wen, CHAI A-li, WANG Wei-wei, LI Bao-ju
    Scientia Agricultura Sinica. 2015, 48(19):  3848-3856.  doi:10.3864/j.issn.0578-1752.2015.19.007
    Abstract ( 481 )   HTML ( 1 )   PDF (1656KB) ( 460 )   Save
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    【Objective】The systemic long-lasting and broad-spectrum resistance of cucumber seedlings was induced with natural or synthetic compounds. The objective of this study is to shed light on the resistance induced by fluoro-substituted benzothiadiazole derivatives (FBT) to cucumber Fusarium wilt and provide data for elucidating its mechanism.【Method】Before transplanting, 50 mg·L-1 of FBT was used to spray so as to induce the cucumber seedlings at 3-4 true leaf stage. Three days later, the cucumber seedlings were transplanted into the soil with Fusarium oxysporum f. sp. cucumerinum Owen, and sprayed with FBT so as to induce resistance for the second time. Sequently, the cucumber seedlings were sprayed every 7 days and for 3 times continuously. Meanwhile, contrast agent BTH also was used with the same induction method and 70% thiophanate-methyl at 1 500 times was applied through root-irrigation. The efficiency on the blight disease resistance was assessed through investigating disease index. For studying the influence of FBT on F. oxysporum f. sp. cucumerinum invasion of cucumber, the cucumber’s radicle length was sprouted to 0.5 cm and then the radicle immersed in 50 mg·L-1 of FBT. The resistance of cucumber seedlings was induced another time at the stage of 2 pieces of leaves. Since then, the cucumber seedlings were sprayed in order to induce the resistance every 7 days and total for 3 times. Twenty-four hours after the last induction, cucumber was inoculated with F. oxysporum f. sp. cucumerinum by the root-drenching method. Treated roots and the controls were harvested at 1, 3, 5, 7, 9, 11, 14, 16, 20, 24 and 29 days after inoculation and then cleaned in ice water. After induction, acid fuchsin dyeing technology was used to evaluate the FBT impacts on fusarium infection. In addition, the Maule reaction and toluidine blue staining technique were used to evaluate the change of lignin and phenolic substances in cucumber root tissue deposition. Moreover, metabolic alterations were investigated and enzyme activities of HRGP and β-1,3-glucanase were determined to analyze the physiological changes in BTH-treated cucumber by speetropho tometer-chromatometry.【Result】Because of the expression of induced resistance, it was found that the control efficiency of FBT was 62.01% at concentration of 50 mg·L-1, higher than BTH treatment and thiophanate-methyl treatment at the same conditions. Seven days after inoculation, colonization of F. oxysporium f. sp. cucumerinum began to appear in the cucumber root tissue induced by FBT. Whereas a large number of hyphae and spores had appeared on the cucumbers not induced with FBT, it indicated that cucumber induced with FBT could gain resistance so as to surpresse the colonization of pathogens. The brown lignin was accumulated in the cucumber roots induced by FBT, especially in epidermal cell walls and phloem xylem. Observation of phenolic substances showed that, two days after FBT induction, phenolic compounds appeared in 4 and 6 days, fluorescence signal in root tissue stronger, phenolic accumulated. Eight days after induction, the phenomena that root tissue fluorescence signal disappeared, phenolic no deposition, lignin and phenolic compounds accumulation increased, resulted in the enhancement of the resistance of the cell wall.【Conclusion】It was found that the resistance induced in cucumber seedlings by FBT includes the accumulation of secondary metabolites in the root and the activation of disease-resistant proteins, suggesting FBT is a promising novel inducer.
    Tissue Specific Expression of Genes Encoding Detoxification Enzymes in the Larvae of Bactrocera dorsalis Under β-Cypermethrin Stress
    SHEN Guang-mao, WANG Xiao-na, HUANG Yong, DOU Wei, WANG Jin-jun
    Scientia Agricultura Sinica. 2015, 48(19):  3857-3865.  doi:10.3864/j.issn.0578-1752.2015.19.008
    Abstract ( 343 )   HTML ( 2 )   PDF (519KB) ( 621 )   Save
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    【Objective】Based on the transcriptome data of Bactrocera dorsalis, the aim of this study is to systematically analyze the expression of genes in larvae encoding detoxification enzymes under β-cypermethrin exposure, identify important detoxification genes, and focus on the discussion of biological significance of tissue specific expression of these genes. 【Method】 The expressions of 30 detoxification genes in the whole body of larvae, midgut, and fat body were detected under β-cypermethrin exposure by using qPCR.【Result】Under the stress of β-cypermethrin, the qPCR results of 8 GSTs genes showed that the mRNA levels of 4 Delta class genes significantly increased. The expressions of genes from Delta class were up-regulated to 5.6-32.5 folds in the midgut and fat body, but the expressions of other GST genes did not show very active reaction when exposed to β-cypermethrin. Meanwhile, although the expressions of Delta class GSTs genes significantly increased in midgut and fat body, the expression changes of these genes were not so apparent in the whole body. As for the 6 CCEs genes, their expressions were up-regulated in some degree in the whole body, and α-E3 was up-regulated to 4.1 folds. However, in the midgut and fat body, the expressions of these genes did not increase significantly. The qPCR results of 16 P450s genes showed that the expressions of most P450s genes were up-regulated, especially the genes from CYP4 and CYP6 families. In CYP4 family, the expressions of CYP4D47 (midgut), CYP4E9 (whole body and fat body), CYP4P5 (whole body and fat body) were highly up-regulated. The expression of CYP4AD1 increased in some degree in the midgut and fat body, but no significant changes were found in the whole body. CYP4S18 and CYP4AC4 did not show any positive reaction or their expression changes were not apparent. In CYP6 family, the expressions of CYP6A48 and CYP6A41 had a significant increase in the fat body, 18.0 and 9.6 folds up-regulated, respectively. The expression of others like CYP6G6 and CYP6A50 also increased. The expression of CYP6EK1 was up-regulated by 4.8 folds in the whole body, but did not change in midgut and fat body. The expressions of the other genes, such as CYP12C2, CYP12N1, CYP9F6, and CYP309B1 also significantly increased in the fat body when exposed to β-cypermethrin. But the expression of CYP317B1 did not change under the stress.【Conclusion】When under the stress of β-cypermethrin, the detoxification genes in B. dorsalis larvae were highly enriched in midgut and fat body, but showed a balance in the whole body. It suggests that when under the toxic stress, the pests may have special strategy for efficient use and proper distribution of energy, which increase the energy efficiency to decrease the extra energy cost to resist the damage of exogenous toxin. This kind of strategy might be an important acclimation of insects to avoid fitness cost when under the environmental pressures.
    SOIL & FERTILIZER·WATER-SAVING IRRIGATION·AGROECOLOGY & ENVIRONMENT
    Survey and Dietary Exposure Assessment of Cadmium in Wheat from Two Main Wheat-Producing Regions in China
    LU Mei-bin, CHEN Zhi-jun, LI Wei-xi, HU Xue-xu, LI Jing-mei, WANG Bu-jun
    Scientia Agricultura Sinica. 2015, 48(19):  3866-3876.  doi:10.3864/j.issn.0578-1752.2015.19.009
    Abstract ( 379 )   HTML ( 8 )   PDF (1150KB) ( 1215 )   Save
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    【Objective】 The objective of this study is to analyze the variation and characteristics of cadmium levels in wheat of two main wheat-producing regions in China and assess the risk degree and law of cadmium in wheat from dietary exposure to different consumer groups. Results of this study could provide a scientific basis for the supervision of quality and safety of agro-products in China.【Method】 A total of 393 wheat samples were collected from two main wheat-producing regions of Huang-Huai-Hai and the middle and lower reaches of the Yangtze River, including 8 provinces (cities). The wheat samples were treated with nitric acid and hydrogen peroxide solution by low-temperature digestion. The cadmium contents of the samples were detected by graphite furnace atomic absorption spectrophotometer (GFAAS). A non-parameter probabilistic model was used to assess the dietary exposure risk based on the information of wheat consumption and body weight of target populations. 【Result】The mean of cadmium contents of wheat in China’s two main wheat-producing regions was (0.032±0.051) mg·kg-1, and cadmium contents in 97.5% wheat samples were below maximum allowable concentration (MAC). The average content of cadmium in wheat was (0.060±0.091) mg·kg-1 in the middle and lower reaches of Yangtze River region, while the level was (0.024±0.025) mg·kg-1 in Huang-Huai-Hai region. The cadmium levels of wheat in the middle and lower reaches of Yangtze River region were significantly higher than those in Huang-Huai-Hai region. There were different cadmium concentrations of wheat between different planting regions. Also it showed difference in the same variety from area to area. Considering the disparity of age and the difference of city and countryside, generally, differences were found in dietary exposure among the various consumer groups all over the country. Different consumption groups with exposure levels of cadmium in wheat had an overweighed ranking for the groups of countryside, younger ages, respectively. Considering the disparity of gender, the exposure levels of cadmium in wheat in females were higher than the levels in males (age<18 years old). When the age >18 years old, the opposite case presented. The percent ratio of the mean of the assessed dietary exposure value to PTMI recommended by JECFA (Joint FAO/WHO Expert Committee on Food Additives, Provisional tolerance monthly intake: 25 μg·kg-1 bw) was in the range of 9.0%-16.8%, the 97.5th percentile level was from 28.9% to 53.8%. The highest exposure level occurred in girls of 4-7 years old, and the lowest was found in women of 60-70 years old. In the city, considering the disparity of age and gender, the different consumption groups with exposure levels of cadmium in wheat have an overweighed ranking for the groups of younger ages and masculinity, respectively. Although the exposure level of females was higher than those of males in the age of 60-70 years old and there were lower exposure levels in the age of 14-18 and 45-60 years old in females in the city. The percent ratio of the mean of the assessed dietary exposure value to PTMI was in the range of 7.3%-13.1%, the 97.5th percentile level was from 23.4% to 42.2%. The highest exposure level occurred in boys of 4-7 years old, and the lowest was found in women of 14-18 years old. The assessed dietary exposure level of rural consumer groups had the similar rules of all over the country, the exposure value in the countryside to PTMI was in the range of 9.6%-17.9%, and the 97.5th percentile level was from 30.9% to 57.4%. The highest exposure level appeared in girls of 4-7 years old, and the lowest was found in women of 60-70 years old. 【Conclusion】 The study showed that the cadmium pollution in wheat was at lower level. There were significant differences in cadmium contents of wheat among the areas and within the varieties. The differences were found in dietary exposure among the various consumer groups. The probability of health risk of cadmium via wheat exposure was at the acceptable level.
    Diagnosing Nitrogen Nutrition Status of Winter Rapeseed via Digital Image Processing Technique
    WEI Quan-quan, LI Lan-tao, REN Tao, WANG Zhen, WANG Shao-hua, LI Xiao-kun, CONG Ri-huan, LU Jian-wei
    Scientia Agricultura Sinica. 2015, 48(19):  3877-3886.  doi:10.3864/j.issn.0578-1752.2015.19.010
    Abstract ( 530 )   HTML ( 12 )   PDF (300KB) ( 600 )   Save
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    【Objective】 In order to provide a scientific basis for digital image processing technique in nitrogen nondestructive diagnosis of winter rapeseed, a field experiment was carried out to explore the feasibility of digital image processing technique in nitrogen nondestructive diagnosis, determine the best digital parameter and regression equation. 【Method】 A field experiment was conducted with different nitrogen application rates (0, 90, 180, 270 and 360 kg·hm-2). The pictures of winter rapeseed canopy were obtained with a digital camera, meanwhile, the conventional nitrogen diagnosis parameters of aboveground biomass, nitrogen concentration in leaf and leaf chlorophyll content at six-leaf period, ten-leaf period, bud period and blooming period, were determined and the correlations were analyzed. 【Result】 The red color intensity (R), normalized redness intensity (NRI) and the ratio of greenness and blueness intensity (G/B) had significant inverse correlations with the conventional N diagnosis parameters. But green color intensity (G), blue color intensity (B), the ratio of greenness and redness intensity (G/R), the ratio of blueness and redness intensity (B/R), the normalized greenness intensity (NGI) and the normalized blueness intensity (NBI) showed a significant positive correlation. Compared with other digital index, the normalized redness intensity (NRI) showed a prominently and very prominently inverse relationship with conventional N parameters and the regression equation with N application rate, aboveground biomass, leaf N concentration, chlorophyll in leaf, N uptake and N nutrition index is y(t·hm-2)=-8.003x+2.706, y(t·hm-2)=-106.072x+38.200, y(g·kg-1)=-692.99x+261.84, y(mg·g-1)=-12.750x+5.665, y(kg·hm-2)=-4087.416x+1414.274 and y=-27.198x+9.812, the correlation values (R2) is 0.917**, 0.746** and 0.953** between measured and estimated leaf N concentration, chlorophyll in leaf and NNI, RMSE and RE were 0.821, 26.32%, 0.330, 28.57% and 0.228, 28.39%, respectively. 【Conclusion】Digital image processing technique can be used in nitrogen nondestructive diagnosis of winter rapeseed, the stage before bud period is crucial period with NRI as the index for the N nitrogen diagnosis using digital image processing technique, and NRI is the best diagnosis index with linear regression equation.
    HORTICULTURE
    Establishment of Molecular ID for Some Apple Germplasm Resources
    GAO Yuan, LIU Feng-zhi, WANG Kun, WANG Da-jiang, GONG Xin, LIU Li-jun
    Scientia Agricultura Sinica. 2015, 48(19):  3887-3898.  doi:10.3864/j.issn.0578-1752.2015.19.011
    Abstract ( 419 )   HTML ( 3 )   PDF (496KB) ( 963 )   Save
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    【Objective】 A total of 131 apple germplasms including landraces, bred cultivars and related species selected from the National Repository of Pear and Apple Germplasm Resources in Xingcheng, China were studied with tailed primer M13 microsatellite markers (TP-M13-SSR). An analysis was made to establish the molecular ID of 131 apple germplasms.【Method】Based on genetic fingerprints, germplasms were distinguished with selected SSR markers, alleles that were amplified by each marker were coded, then was combined as a molecular ID.【Result】Two accessions selected from a total of 131 accessions were used for optimization of the first PCR detecting conditions and SSR primer screening. 16 pairs of TP-M13-SSR primers with high stability and good repeatability were used to establish the fingerprints of 131 accessions of Malus Mill.. By using 16 selected SSR markers, 326 polymorphic sites were detected with a mean value of 20.3. The amplification of CH05d04 for all germplasms obtained 49 alleles with the highest expected heterozygosity 0.878, and the amplification of CH01f07a for all germplasms obtained 48 alleles. The polymorphism information content was calculated by PopGen32 with 0.7558 on average. 16 SSR markers could distinguish 11 accessions at least, 71 accessions at most, and 49 accessions on average. The identified rate was 8.09%-52.21%. The identified rate of CH01f07a was the highest, and the identified rate of BGT23b was the lowest. Based on the PIC of amplification and identified rate, each parir of two SSR primers was combined together to identify all the accessions. The combination of CH04h02 and CH01f07a could distinguish 120 accessions at most. More SSR primers were combined together. Finally three SSR primers could distinguish all the accessions. All the alleles of the three core SSR markers were sequenced from small to large, and the assignment was from number 01. A character string was constituted by combining all the codes of the three primers for every accession. By using barcode technology molecular ID can be transferred into a barcode ID that can be quickly scanned by machine.【Conclusion】Based on the PIC of amplification and identified rate, core SSR primers can be screened out to distinguish all the landraces, bred cultivars, and related species. And by constructing fingerprints, every apple germplasm obtains its differentiable molecular ID that can be recognized by the machine. The purpose was to distinguish the most apple germplasms by using the least and the most specific primers.
    Oil Body Observation in Seed Development and Its Analysis in Seed of Juglans regia ‘Wen185’ and J. regia ‘Xinxin2’ in Period of Seed Maturity
    CHEN Hong, PAN Cun-de, WANG Bei, XIAO Zhen-zhen, HU Yuan, HU Guo-jun
    Scientia Agricultura Sinica. 2015, 48(19):  3899-3909.  doi:10.3864/j.issn.0578-1752.2015.19.012
    Abstract ( 375 )   HTML ( 6 )   PDF (3021KB) ( 707 )   Save
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    【Objective】Through the observations of different varieties of walnut seed oil body development processes and the analyses of the mature cotyledon oil body characteristics, we aimed at finding walnut seed oil body synthesis initiation time and location, understanding the relationship between the oil rate and the differences in the cotyledon oil of the seeds of different varieties and deepen the understanding of Xinjiang early fruit walnut seed cell ultrastructure.【Method】The oil content were tested by a fat analyzer. The ultrathin sections of resin embedded seeds were observed by a Transmission Electron Microscope (TEM). 【Result】 Juglans regia ‘Wen185’ and J. regia ‘Xinxin2’seeds were used as materials. The results showed that, in the embryo, the cell can be distinguished from each other and organelles such as vacuole, nucleolus, chloroplast, starch grain, endoplasmic reticulum, and mitochondrion, the Golgi’s body appeared in each cell after 30 days after flowering (DAF). The number of endoplasmic reticulum were increased after 50 DAF. The tip of the double membrane of endoplasmic reticulum began to be inflated and oil bodies were observed after 60 DAF. The number of oil bodies increased gradually until the fruit matured. The number of endoplasmic reticulum decreased after 90 DAF and organelles disappeared after 110 DAF. The period of organelles which were observed in the cotyledon was later than the embryo. In the period of seed maturity, cells consisted of oil bodies and protein bodies in the cotyledon. The sizes and shapes of the oil bodies were different. Most of oil bodies showed an elliptical or irregular shape. The J. regia ‘Wen185’ had more oil content than J. regia ‘Xinxin2’. Also, it was found that the mean size of the oil bodies of the cotyledons for J. regia ‘Wen185’ was significantly different from the oil bodies of the cotyledons for J. regia ‘Xinxin2’. J. regia ‘Wen185’ has less number of oil bodies per cell than J. regia ‘Xinxin2’. J. regia ‘Wen185’ has larger oil bodies than J. regia ‘Xinxin2’. 【Conclusion】These results provide that oil bodies were originated from the endoplasmic reticulum. The period of organelles which were observed in the cotyledon was later than in the embryo. The length and total sum of the areas of the oil bodies correlate well with the oil content of walnut of Juglans regia ‘Wen185’ and J. regia ‘Xinxin2’.
    ANIMAL SCIENCE·VETERINARY SCIENCERE·SOURCE INSECT
    Effects of Saccharomyces cerevisiae on the Expression of SBD-1 in Cultured Ruminal Epithelial Cells of Sheep
    JIN Xin, ZHANG Man, FAN Yan-ru, WANG Pei, YANG Yin-feng
    Scientia Agricultura Sinica. 2015, 48(19):  3910-3918.  doi:10.3864/j.issn.0578-1752.2015.19.013
    Abstract ( 321 )   HTML ( 5 )   PDF (2282KB) ( 486 )   Save
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    【Objective】The regulatory effect of probiotic Saccharomyces cerevisiae on the β-defensin-1 (Sheep Beta-Defensin-1, SBD-1) expression was explored in cultured ruminal epithelial cells of sheep, which would provide a theoretical foundation and basis for the regulation mechanism of probiotics to defensins at the molecular level. 【Method】 Firstly, the primary culture of the sheep ruminal epithelial cells were carried out after treating the sheep ruminal tissue with PBS and antibiotic. When the number of cells in primary cell culture bottles grew to more than 85%, the cells were passaged to 12-well plates for cell stimulation test, meanwhile, a parallel plate count test was conducted for five times after the activated and purified S. cerevisiae were cultured at 25℃, 180 r/min aerobically for 48 h. A concentration of 5.2 × 108 CFU·mL-1 S. cerevisiae was obtained according to processing method of plate count, then the concentration was further diluted to the concentrations of 5.2×107, 5.2×106, 5.2×105 and 5.2×104 CFU·mL-1 by a serial dilution. The cultured sheep ruminal epithelial cells were stimulated for different times (2, 4, 8, 12 and 24 h) by different concentrations of S. cerevisiae (5.2×104 - 5.2×108 CFU·mL-1), and a control group was set up with DMEM/F12 medium instead of S. cerevisiae. Secondly, the total RNA was extracted from the tested cells and reversely transcripted into cDNA, then different expressions of SBD-1 were examined by real time fluorescence quantitative PCR and enzyme-linked immunosorbent assay (ELISA) tests. Finally, the related differences analysis of data was done by SAS 9.2 software. 【Result】The realtime fluorescence quantitative PCR results showed that the expression of SBD-1 mRNA in each time group increased and then decreased following the increase of concentrations, the expression of SBD-1 mRNA was the highest when the cells were stimulated with the concentration of 5.2 × 107 CFU·mL-1, and significantly higher than the control group and the other concentration groups (P<0.01). When the concentration was constant, the expression of SBD-1 mRNA increased with the extension of stimulating time, which the expression of SBD-1 mRNA was the highest at 12 h, then downward. Therefore, the expression of SBD-1 was the highest at the concentration of 5.2 × 107 CFU·mL-1 and 12 h stimulating, compared with the other time groups at this concentration was significantly different (P<0.01 ). The results of ELISA test showed that the expression trend of SBD-1 protein was consistent with the SBD-1 mRNA. After the sheep ruminal epithelial cells were stimulated with different concentrations of S. cerevisiae for 2, 4, 8, 12 and 24 h, the expression of SBD-1 protein in the co-culture supernatants increased, and there was a significant difference compared with the control group (P<0.01 ). The expression of SBD-1 protein reached the highest level at concentration of 5.2 × 107 CFU·mL-1 and 12 h stimulating, the difference was significant compared with the each time group under this concentration (P<0.01).【Conclusion】The results of this study indicated that S. cerevisiae could improve the expression of SBD-1 in ruminal epithelial cells of sheep. And the level of SBD-1 expression was the highest when the ruminal epithelial cells were stimulated for 12 h with the concentration of 5.2 × 107 CFU·mL-1.
    Polymorphism of Pre-microRNA-1658 Gene in Chicken
    GENG Li-ying, ZHANG Chuan-sheng, ZHAO Shu-yu, CHEN Juan, GONG Yuan-fang, LIU Zheng-zhu, ZHU Wen-jin, LI Xiang-long
    Scientia Agricultura Sinica. 2015, 48(19):  3919-3930.  doi:10.3864/j.issn.0578-1752.2015.19.014
    Abstract ( 303 )   HTML ( 1 )   PDF (2604KB) ( 456 )   Save
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    【Objective】 To investigate chicken gga-mir-1658 precursor area gene genetic variations/haplotypes and its distribution among all varieties. To analyze its effect on microRNA secondary stem-loop structure and target gene selection. The purpose was to screen the mutation sites as having potential biological function, which might lay the foundation to further reveal its influence on gga-mir-1658 gene expression regulation and phenotypic effects. 【Method】 A pair of specific primers were designed according to the chicken gga-mir-1658 genome sequence (GenBank accession number: NR_035151.1). The polymorphism detection was performed on gga-mir-1658 gene precursor area of 220 individuals including three chicken breeds: Taihang chicken (95), Beijing Fatty Chicken (83) and Leghorn chicken (42) by the PCR products direct sequencing. The comparison analysis and secondary structural simulation of gga-mir-1658 genome sequences were performed using DNAman, MEGA and mfold software. The matching linkage disequilibrium analysis and haplotype analysis were performed by SHEsis and Haploview software. The predictive parsing on the free energy changes of gga-mir-1658 target genes and their complexes was performed by miRanda software. 【Result】There were 6 mutation sites in pre-gga-mir-1658 gene. Among them, secondary allele frequency ≥5% included g.28 C>G, g.31 C>T, g.70G>A and g.71G>-. Four variable sites were positioned in the seed zone of gga-mir-1658 gene mature body. The genetic variation analysis showed that g.70G>A site manifested low polymorphism (PIC<0.25), and the other three sites manifested moderate polymorphism (0.25<PIC<0.50). The compatibility test showed that the other variation sites were in a Hardy-Weinberg equilibrium state in all breeds except for Leghorn chicken g.31 C>T, g.71G>-, Taihang chicken g.71G>- and Beijing Fatty Chicken g.70G>A site (P>0.05). Linkage disequilibrium and haplotype analysis showed that there was a weak linkage equilibrium between mutated sites; 11 haplotypes were detected among the three breeds. Of which, H1 (C C G -) and H11 (G T G G) were the dominant haplotypes. The frequencies were greater than 25%. Bioinformatics analysis showed that the mutation in the seed zone could influence the space configuration and free energy of gga-mir-1658 gene precursor secondary structure. Of which, H6 haplotype mutants was highest (41.00 kcal·mol-1). H2 and H5 haplotype mutants were lowest (35.70 kcal·mol-1); The predominant haplotype H1 and H11 mutants were -36.10 kcal·mol-1 and 40.04 kcal·mol-1. Different haplotype mature seed zone sequences of gga-mir-1658 gene were different. The gga-mir-1658-5p included two kinds of seed sequences: “AUACCAU” and “AUACCAC”. The gga-mir-1658-3p included four seed zone sequences: “AACUCUG”, “AGCUGUG”, “AACUGUG” and “AGCUCUG”. The bioinformatics analysis of gga-mir-1658 predictable target gene showed that seed zone could affect the selection of gga-mir-1658 mature body on the target gene. They were mainly enriched in gene expression regulation, cell apoptosis, immune system development and B cell activation, other basic biological processes. 【Conclusion】 (1) There were four mutation sites having potential biological function and phenotypic effects in gga-mir-1658 gene seed zone. They could constitute 11 haplotypes. Of which, H1 (C C G -) and H11 (G T G G) were the dominant haplotypes in Beijing Fatty Chicken, Taihang chicken and Leghorn chicken. (2) The seed zone mutation could affect the stability of gga-mir-1658 gene precursor secondary structure and the selection of target genes, which might be the important functional site having potential phenotype effect.
    Establishment of a Detection Method for the Immune Colloidal Gold of Phenylethanolamine A in Animal Urine
    NIE Wen-ying, LUO Xiao-qin, LI Jin-chao, LI Hang, WU Guang, WANG Lin-chen
    Scientia Agricultura Sinica. 2015, 48(19):  3931-3940.  doi:10.3864/j.issn.0578-1752.2015.19.015
    Abstract ( 338 )   HTML ( 1 )   PDF (1192KB) ( 638 )   Save
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    【Objective】The aim of the study is to establish a detection method for the immune colloidal gold of phenylethanolamine A in animal urine. 【Method】Phenylethanolamine A hapten was synthetized by nucleophilic substitution with phenylethanolamine A and bromo aldehyde. Phenylethanolamine A hapten was coupled with BSA and OVA, respectively, to prepare immunogen and envelope antigen. The coupling ratio of hapten-BSA/OVA was determined by TNBS. Monoclonal antibody was acquired from mice which were immunized with hapten-BSA. The indirect ELISA method has been established for sensitivity and specificity detection of monoclonal antibody. A total of 13 compounds whose structures were similar with phenylethanolamine A were used to test the specificity and calculate the cross reaction rate of monoclonal antibody. A rapid detection method for immune colloidal gold of phenylethanolamine A in animal urine was established by the parts of strip prepared and assembled. The strip’s detection limits, false positive rate and false negative rate were determined. Compared the immunochromatography assay (ICA) with ELISA whose limited ranges of standards’ concentration were 0, 0.1, 0.3, 0.9, 2.7 and 8.1 μg·L-1, the detection limits for pig urine was 0.5 μg·L-1 and the ranges of percent recovery were 75%-105%.【Result】Phenylethanolamine A derivative was prepared that amino terminal of phenylethanolamine A connected with the connecting arm whose aldehyde group was on the end. Phenylethanolamine A hapten connected with BSA and OVA successfully by TNBS and the coupling ratio between hapten with BSA and OVA were 11.38 and 10.66, respectively. The titer and ascites of MC-73 hybridoma cell strain were 1﹕2×103 and 1﹕5×107 which were higher than MC-33 and MC-29. IC50 of monoclonal antibody against phenylethanolamine A was 0.365 μg·L-1 which was more sensitive than MC-33 and MC-29. The cross reaction rate with 13 compounds whose structures were similar with phenylethanolamine A were less than 1%. The detection limit, false positive rate and false negative rate were 5 μg·L-1, 0 and 0, respectively. Detection results of phenylethanolamine A in pig urine between ICA and ELISA were the same.【Conclusion】The phenylethanolamine A strip with high sensitivity, specificity, accuracy and rapidity was developed in this study which would be used to test animal urine in 5 min without pre-treatment of urine samples. It would be efficient for mass samples screening in primary laboratory and enterprises.
    RESEARCH NOTES
    Genetic Analysis of New Wheat Variety Zhoumai 23 and Screening of Specific Primers
    ZOU Shao-kui, YIN Gui-hong, TANG Jian-wei, HAN Yu-lin, LI Nan-nan, LI Shun-cheng, HUANG Feng, WANG Li-na, ZHANG Qian, GAO Yan
    Scientia Agricultura Sinica. 2015, 48(19):  3941-3951.  doi:10.3864/j.issn.0578-1752.2015.19.016
    Abstract ( 490 )   HTML ( 7 )   PDF (632KB) ( 603 )   Save
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    【Objective】 The objective of this study was to understand the genetic contributions of two parents to wheat variety Zhoumai 23 and the genetic composition of Zhoumai 23 in the Huang-Huai region, and the specific primers screened for the detection of the varietal authenticity of Zhoumai 23.【Method】 A total of 340 SSR markers covering 21 wheat chromosomes were used in a simple sequence repeat (SSR) markers analysis. Parsed the transfer frequency and genetic contribution rate of parental genetic material to Zhoumai 23. This also could be screened for several primers of Zhoumai 23 different from either parent, further screening of these specific markers using sister lines, derived varieties of Zhoumai 23. Finally, with the selected 1-2 specific primers of Zhoumai 23, and confirmed final screening of specific primers using Zhoumai 22, Jimai 22, Aikang 58, Zhengmai 366, a total of 14 materials of the main push varieties in the Huanghuai wheat region.【Result】 The results indicated that there were large differences in the genetic contribution from double parents to Zhoumai 23, the rate of the genetic contribution from Zhoumai 13 was 63.04%, much higher than that (36.96%) from Xinmai 9. Partial segregation occurred during the breeding of the parent material in the Zhoumai 23. The genetic contribution rate changes greatly in the different genomes and the chromosome level. The genetic contribution range from Zhoumai 13 to Zhoumai 23 was 23.1% (1B)-100% (4A, 6A, 3B, 4B, 6B, 4D) and from Xinmai 9 to Zhoumai 23 was 0 (4A, 6A, 3B, 4B, 6B, 4D)-76.9% (1B). The study identified seven specific sites in Zhoumai 23 from 147 polymorphic markers, named Xwmc344, Xbarc84, Xwmc326, Xwmc468, Xwmc479, Xgwm428 and Xcwm65. One specific primer, Xcwm65, can be used to differentiate Zhoumai 23 from the main varieties in Huang-Huai wheat region. Sublines A3, A4, A5 and derived varieties B7, B8, B12, and B15 of Zhoumai 23 were still difficult to distinguish. Therefore, it is needed to identify the different molecular markers to distinguish between Zhoumai 23 sister lines and its derivatives. 【Conclusion】 The study revealed a genetic contribution from the parents to Zhoumai 23, master of Zhoumai 23 genetic composition and the mapping of the gene map, and found a specific primer for the identified authenticity of Zhoumai 23.
    Study on Yield Loss of Summer Maize Due to Lodging at the Big Flare Stage and Grain Filling Stage
    LI Shu-yan, MA Wei, PENG Ji-yong, CHEN Zhong-min
    Scientia Agricultura Sinica. 2015, 48(19):  3952-3964.  doi:10.3864/j.issn.0578-1752.2015.19.017
    Abstract ( 558 )   HTML ( 16 )   PDF (572KB) ( 658 )   Save
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    【Objective】The objective of this study was to analyze the effects of lodging on dry matter, yield components and final yield of summer maize, and further to evaluate the yield losses due to different types of lodging at different growing stages. 【Method】The field experiments were conducted during two growing seasons of summer maize with the variety of Xundan 20 from 2011 to 2012 at Zhengzhou Agro-meteorological Experiment Station. The lodging was implemented by kicking down the maize plants artificially at two growing stage, the big flare stage (B) and the grain filling stage (F). And at each stage, the lodging included four types, light lodging of roots (R1, in which the angle between the lodged stem and ground was between 30 and 60 degree), serious lodging of roots (R2, in which the angle between the lodged stem and ground was lower than 30 degree), lower node lodging of stem (SL, in which the bend position was below the third node) and upper node lodging of stem (SH, in which the bend position was above the third node). LAI, dry matter, yield and yield components were measured during experiments, and the relative reduction percentage was calculated which indicating the damage degree.【Result】The results showed that LAI was significantly decreased by lodging. The relative reduction percentage was larger in two treatments of stem lodging at the filling stage, with the value of 38.9% (FSL) and 50.7% (FSH), respectively. The accumulation of aboveground dry matter was significantly reduced after lodging (P<0.05) in all treatment except FR1, and the reduction percentages were larger in two treatments of stem lodging at the filling stage, with the value of 34.9% (FSL) and 46.8% (FSH), respectively. Lodging also affected the partition of dry matter among leaves, stems and corncobs, with the increased partition to leaves and stems but the decreased partition to corncobs. In two treatments of stem lodging at the filling stage, the weight of leaves and stems was 9.2% (FSL) and 3.7% (FSH) higher than CK, but the weight of corncobs was 9.9% and 7.0% lower than CK. In addition, lodging made the corncobs shorter and thinner, and with longer barren ear tips. The length of corncobs in treatments with lodging at the filling stage was 3 to 4 cm shorter compared to CK. Except for the FSL treatment in 2012, the rate of barren ear tips in treatments with lodging at the filling stage were statistically and significantly higher than CK, but it was not significant with treatment with lodging at the big flare stage. And the rate of barren ear tips was the highest in FSH treatment in 2011, with the percentage of 27.4%. Lodging caused the decrease of both grains per ear and 100-grain weight. Due to the differences of experimental conditions between two years, results were slightly different. In 2011, grains per ear were significantly less than CK in all treatments, but in 2012, they were only significant in the treatments with stem lodging at filling stage. The effects of lodging on 100-grain weight were more significant in treatments with lodging at filling stage. Lodging also significantly decreased the final grain yield. According to the analysis of yield components, the yield reduction by lodging was due to the decreases in both grains per ear and 100-grain weight. And the correlation coefficients of the two components with grain yield were 0.729 and 0.842 (P<0.01), respectively. Except for BR1, the final yields in all treatments were significantly lower than CK. For the lodging at the big flare stage, the yield loss was 13.9%, 27.9% and 27.1% in the treatments of BR2, BSL and BSH, respectively. For the lodging at the grain filling stage, the yield loss was 29.0%, 38.4%, 45.0% and 48.3%, respectively.【Conclusion】Comprehensively, compared between growing stages with the same lodging type, lodging at grain filling stage had more negative effects than that at big flare stage. While compared between lodging types at the same stage, stem lodging had more significant effects than root lodging. There was no obvious difference between treatments with lodging at high node and low node. For yield loss, it was the largest for stem lodging at grain filling stage, and the lowest for root lodging at big flare stage.
    Evolution of Volatile Compounds During the Berry Development of ‘Ruidu Xiangyu’ Grape
    ZHANG Ke-kun, WANG Hai-bo, WANG Xiao-di, SHI Xiang-bin, WANG Bao-liang, ZHENG Xiao-cui, LIU Feng-zhi
    Scientia Agricultura Sinica. 2015, 48(19):  3965-3978.  doi:10.3864/j.issn.0578-1752.2015.19.018
    Abstract ( 468 )   HTML ( 3 )   PDF (556KB) ( 585 )   Save
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    【Objective】 Table grapes with early maturity and fragrant flavors play an important role in fruit industry and are more attractive to consumers when compared with other cultivars. Comprehensive study on the evolution of volatile compounds of early maturing variety is significant for breeding and evaluation of new cultivation technique.【Method】This study used headspace solid phase micro-extraction method and the technology of gas chromatography-mass spectrometry to analyze the changes of volatile compounds and related metabolic pathways of grape berries (V. vinifera cv. Ruidu Xiangyu), which were sampled from 3 weeks to 12 weeks past blossom. In the meantime, the method of PCA was used to clear the relationship between developmental stages and volatile compounds. 【Result】 ‘Ruidu Xiangyu’ came into the stage of verasion at 5 weeks after blossom and into the stage of maturation at 9 weeks after blossom. The fragrant monoterpenes like linalool, nerol, geraniol, citronellol and terpineol accumulated differently at the full developmental process. The maximum content of nerol and geraniol showed up at the beginning of maturation while linalool and terpineol in the middle of it. And the content of citronellol changed irregularly at maturation. E-2-hexenal and hexanal are the main C6 compounds at maturation, and their content went to the peak after verasion and then decreased. Z-3-hexenal mainly existed at young fruit period, of which the content was low at verasion. Additionally, the content of norisoprenoids like β-lonone, damascenone and aromatic compounds like benzaldehyde was very low at maturation. Considering the content change of various compounds, terpenes accumulated differently at different developmental stages and decreased at maturation. Esters mainly accumulated at young fruit period and postveraison stage while aldehydes and ketones were evident at preveraison stage. C6 compounds increased during veraison and then decreased with alcohols. Aromatic compounds and volatile acid accumulated at preveraison and postveraison stage respectively. The key stages for content change are also different to various volatile compounds. Verasion is the key stage for aldehydes and ketones, while major esters and aromatic compounds change before it, and terpenes, C6 compounds and volatile acid change after it. 【Conclusion】Our study indicates that the Z-3-hexenal route from splitting linolenic acid is active at early stage. Other Lipoxygenase (LOX) routes and synthesis of terpenes become active during veraison while accumulations of most compounds become weak at maturity. The key stage for the formation of its characteristic aroma is from verasion to maturity. Major fragrant terpenes accumulate differently in ‘Ruidu Xiangyu’ grape and linalool and α-terpineol decrease later than other monoterpenes. Considering C6 compounds and terpenes, properly delayed harvest will be better for the realization of its fragrance.
    Apple Pomace Dietary Fibre Modification by Hydrogen Peroxide
    GENG Yi-wen, HA Yi-ming, JIN Jing, LI Qing-peng
    Scientia Agricultura Sinica. 2015, 48(19):  3979-3988.  doi:10.3864/j.issn.0578-1752.2015.19.019
    Abstract ( 328 )   HTML ( 1 )   PDF (1234KB) ( 508 )   Save
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    【Objective】The aim of this study was to explore the effect of hydrogen peroxide modification on the dietary fibre in apple pomace, and provide a simple, effective and cheap method to increase the content of soluble dietary fibre and improve physicochemical properties of apple pomace. 【Method】Different pHs and concentrations of hydrogen peroxide were used to treat the apple pomace which obtained from juice factory. After precipitating by ethanol, drying and grinding, the hydrogen peroxide modified apple pomace was obtained. The physicochemical and structural properties of apple pomace were studied, such as yield, total dietary fibre (TDF) content, insoluble dietary fibre (IDF) content, soluble dietary fibre (SDF) content, water holding capacity, swelling capacity, oil holding capacity, bulk density, colour, thermostability and microstructure. Hydrogen peroxide residue in modified apple pomace was also determined. 【Result】pH of hydrogen peroxide solution could significantly impact the physicochemical and structural properties of apple pomace. In treatment with the same concentration of the hydrogen peroxide, TDF content, water holding capacity, swelling capacity, oil holding capacity of apple pomace were increased in different degrees by acidic (pH 3.8) or neutral (pH 7) hydrogen peroxide treatments, however, there was no significant change in the content of SDF and bulk density. The colour of apple pomace became dark. In alkaline (pH 11.5) hydrogen peroxide treatment, the content of SDF was significantly increased, the water holding capacity, swelling capacity and colour were also improved significantly, the bulk density and TDF content were increased compared with the untreated apple pomace. Through analysis of the thermostability and microstructure, there was no significant difference between modified and untreated apple pomace in thermostability and microstructure in treatments with acidic or neutral hydrogen peroxide, while the thermal property became less stable and the structure became dense and smooth in treatment with alkaline hydrogen peroxide. The concentration of hydrogen peroxide solution could also significantly impact the physicochemical and structural properties of apple pomace. In alkaline conditions (pH 11.5), when using 0% hydrogen peroxide solution treatment, the physicochemical and structural properties of apple pomace were similar to the acidic and neutral hydrogen peroxide treated products. With the increase of hydrogen peroxide concentration, the SDF content was increased from 3.30% to 19.02%-28.32%, increased by 476%-758%, the swelling capacity and the colour of apple pomace were improved, the bulk density were also increased, the water holding capacity was rised firstly and then decreased, the yield, TDF content, and IDF content were decreased, but oil holding capacity was not improved. Moreover, the structural properties of apple pomace were also changed. The thermostability was reduced and the structure became more fragmented with the increase of hydrogen peroxide concentration. The result of hydrogen peroxide residue showed that hydrogen peroxide could be removed completely during the process of treatment, without any residue. 【Conclusion】Alkaline hydrogen peroxide modification could be used as a clean and effective method to increase the content of soluble dietary fibre in apple pomace and improve the physicochemical properties of apple pomace. The effect of modification was closely related to pH and concentration of hydrogen peroxide.