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    16 May 2015, Volume 48 Issue 10
    CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS
    Gene Mapping and Candidate Gene Analysis of a dwarf and deformed flower 2 (ddf2) Mutant in Rice (Oryza sativa)
    ZHANG Ling, GUO Shuang, WANG Ling, ZHANG Tian-quan, ZHUANG Hui, LONG Yu-chen, HE Guang-hua, LI Yun-feng
    Scientia Agricultura Sinica. 2015, 48(10):  1873-1881.  doi:10.3864/j.issn.0578-1752.2015.10.001
    Abstract ( 529 )   HTML ( 3 )   PDF (7472KB) ( 439 )   Save
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    【Objective】A rice mutant, dwarf and deformed flower 2 (ddf2) , was characterized in the present paper. These results will provide a foundation for map-based cloning and functional analysis of the DDF2 gene. 【Method】 The ddf2 mutant was derived from tissue culture mutation generations of indica restorer 602. At the heading stage, the plant height, panicle length, width and length of leaves and internodes of the ddf2 mutant and wild type were measured. Moreover, the phenotype and histology analyses of stem, leaf and spikelet were conducted. A population containing 1 024 F2 mutants from the zhonghua11/ddf2 cross was used to map the DDF2 gene. The expression of candidate genes were detected by real time PCR. 【Result】 The leaves and internodes of ddf2 mutant were all much shorter and thinner than those of the wild type, and the panicleof ddf2 mutant was shorter than that of the wild type. According to the histology analysis, the width between two large veins of leaf was decreased dramatically in ddf2 mutant, while no obvious difference in the numbers of large and small veins was observed between the ddf2 mutant and the wild type. Further analysis showed that the size and number of mesophyll cells in ddf2 leaves were all reduced significantly. Furthermore, parenchyma cells were significantly reduced in size and number in ddf2 stems, while there was no difference in the number of vascular bundles in stem between the ddf2 mutant and the wild type. These results indicated both the cell division and cell expansion were inhibited in ddf2 leaves and stems. Additionally, the spikelet and floral organs in ddf2 mutant display serious defects. In ddf2 florets, the lemmas were distorted while the paleas were degraded in whorl 1. The stamens were degenerated seriously or transformed into pistil-like organs in whorl 3. Elongated sterile lemma and extra florets were often observed in some ddf2 spikelets. Genetic analysis indicated that ddf2 was a nuclear recessive gene. By using 1 024 F2 mutants from the zhonghua11/ddf2 cross, the DDF2 gene was mapped between In/Del markers S-11 and S-14 on the short arm of chromosome 11 near the centromere, with genetic distance of 0.049 cM and 0.098 cM, respectively, and an approximate physical distance of 90.295 kb. The gene was co-segregating with another In/Del marker S-24. There were 12 MSU annotation genes within the mapping region. Based on the qRT-RCR, it was found that the expression of LOC_Os11g17600, encoding an exocyst complex component Sec3_C protein, was down regulated in the ddf2 mutant. Then, the LOC_Os11g17600 was considered as candidate gene of DDF2.【Conclusion】DDF2 is considered to be a novel gene controlling both stem, leaf and floral organ development.
    Functional Analysis of COI1 Genes in Oilseed Rape (Brassica napus L.)
    WANG Wen-jing, YANG Xiao-chuan, DING Yong-qiang, YIN Guo-ying, MA Hao-ran, ZHANG Jie, SHI Xiao-yu, ZHANG Ding-yu, LI Jia-na, ZHANG Hong-bo
    Scientia Agricultura Sinica. 2015, 48(10):  1882-1891.  doi:10.3864/j.issn.0578-1752.2015.10.002
    Abstract ( 477 )   HTML ( 4 )   PDF (2838KB) ( 549 )   Save
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    【Objective】 CORONATINE INSENSITIVE 1 (COI1) is a critical component of jasmonate (JA) receptor complex. The objective of this study is to investigate the spatial expression pattern and regulatory roles of COI1 gene in oilseed rape(Brassica napus), a staple oil crop in the world. 【Method】 The COI1 genes in oilseed rape(B. napus) and its parental species B. rapa and B. oleracea were analyzed based on the genomic data. The spatial transcription pattern of COI1 in oilseed rape was analyzed by RT-PCR with specific primers according to the conserved region of COI1 gene homologs. Then, a cDNA fragment of the conserved region was cloned into vector pTRV2 of the tobacco rattle virus-based VIGS (virus-induced gene silencing) system. And, VIGS techniques were applied to silence COI1 genes in oilseed rape. The rape plants shown to have COI1 genes silenced were used to investigate male fertility and aphid resistance. 【Result】 Analysis of the genomic data of B. rapa, B. oleracea, and B. napus revealed that the genomes of B. rapa and B. oleracea contained 7 highly homologous COI1 genes, which could be classified into 4 subgroups, i.e., COI1a, COI1b.1, COI1b.2, and COI1c, while 8 COI1 genes were presented in the genome of B. napus. Transcriptional assay revealed that the expression of COI1 in oilseed rape is tissue specific. A 505bp fragment of the conserved region of COI1 was introduced into the VIGS vector pTRV2 to develop COI1-silenced plants via VIGS techniques. Twenty-five plants with the transcripts of COI1 down-regulated by over 70% were identified by transcriptional analyses, from which the ten plants with the lowest expression level of COI1 were selected to investigate male fertility and aphid resistance. The results showed that the fertility of COI1-silenced plants was extremely impaired, with no seeds in their siliques. Furthermore, the filaments of COI1-silenced plants were shorter than those of control plants, and over 80% pollens of the COI1-silenced plants were irregular in shape. Aphid resistance assay showed that aphids grew and reproduced much faster on the COI1-silenced plants than on the control plants cultured under the same condition. 【Conclusion】 This study revealed that the expression of COI1 gene in oilseed rape is tissue specific. VIGS-induced silencing of COI1 greatly impaired the male fertility and decreased the aphid resistance of oilseed rape. This work has provided an important start point to dissect the mechanism of JA signaling in oilseed rape.
    Research of Exogenous Hormone Sensitivity on Plant Height of a Dwarf Germplasm in Jute (Corchorus capsularis L.)
    ZHANG Li-wu, HUANG Wen-xuan, TAO Ai-fen, LIN Li-hui, XU Jian-tang, FANG Ping-ping, QI Jian-min
    Scientia Agricultura Sinica. 2015, 48(10):  1892-1899.  doi:10.3864/j.issn.0578-1752.2015.10.003
    Abstract ( 400 )   HTML ( 5 )   PDF (822KB) ( 320 )   Save
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    【Objective】The aim of this study was to illuminate the sensibility of the exogenous hormones to plant height of a dwarf germplasm Aidianyehuangma so as to determine which kind of hormone sensitivity this germplasm belongs to, which will facilitate dissecting the relationship between the genetic basis of plant height and hormonal regulation in jute. 【Method】 Firstly, the performance of important agricultural traits, e.g. plant height, were compared between Aidianyehuangma and the variety Huangma 179 with normal plant height. Secondly, the dwarf germplasm of Aidianyehuangma was treated with different concentrations of exogenous GA3, IAA and BR. And the hypocotyl length in the Ms culture media and plant height variation in the field were investigated. Meanwhile, the hypocotyl length of Huangma 179 and Aidianyehuangma on the GA3 culture media was evaluated. Thirdly, the effects of continuous spraying of 100 mg·L-1 GA3 every 3 days on plant height of Aidianyehuangma were analyzed. Furthermore, the activities of α - amylase of Aidianyehuangma were analyzed after it was once treated with GA3. 【Result】 The comparison analysis of important agricultural traits showed that the plant height of Aidianyehuangma (1.89 m) was significantly lower than that of Huangma 179 (4.28 m) without exogenous hormone treatment, indicating that Aidianyehuangma is a typical dwarf germplasm in jute. After being treated with MS culture media of 0.1 mg.L-1 exogenous IAA and BR, the hypocotyl length of Aidianyehuangma did not show a significant difference with that of the control. However, after being treated with MS culture media of 0.1 mg·L-1 exogenous GA3, the hypocotyl length of Aidianyehuangma showed a significant difference compared with Aidianyehuangma without being treated with GA3, and the hypocotyl length of Aidianyehuangma and Huangma 179 was similar to that of Huangma 179 without being treated with GA3. It suggested that the plant height of Aidianyehuangma is sensitive to exogenous GA3 while that of Huangma 179 is not sensitive to the exogenous GA3. After being treated with different concentrations of exogenous hormone in the field, plant height of Aidianyehuangma with high concentration (100 mg·L-1) of GA3 was significantly higher than that with the other two lower concentrations of GA3. By contrast, plant height of Aidianyehuangma treated with different concentrations of exogenous IAA and BR did not show a significant difference with that of the control. It further suggested that Aidianyehuangma is a GA3 sensitive dwarf germplasm. After being treated with 100 mg·L-1 exogenous GA3 continuously in the field, the plant height of Aidianyehuangma showed a significant difference in comparison of Aidianyehuangma without being treated with GA3, reaching a significant level, and appoached that of Huangma 179. It showed that plant height of Aidianyehuangma could be resumed using exogenousGA3. After Aidianyehuangma was once treated with GA3, α- amylase activity of Aidianyehuangma in the first 3 days began to increase in comparison with that before treatment, and enhanced significantly in the first 6 days, which indicated that the exogenous GA3 could increase the α- amylase activity of Aidianyehuangma. 【Conclusion】Aidianyehuangma is a typical dwarf germplasm in jute. The plant height of Aidianyehuangma is not sensitive to exogenous IAA and BR, but sensitive to the exogenous GA3, and can be resumed to normal plant height with continuous exogenous GA3 treatment. The exogenous GA3 could increase the α- amylase activity of Aidianyehuangma. Aidianyehuangma is a GA3 sensitive dwarf germplasm.
    TILLAGE & CULTIVATION·PHYSIOLOGY & BIOCHEMISTRY·AGRICULTURE INFORMATION TECHNOLOGY
    Recent Progresses in Research of Crop Patterns Mapping by Using Remote Sensing
    HU Qiong, WU Wen-bin, SONG Qian, YU Qiang-yi, YANG Peng, TANG Hua-jun
    Scientia Agricultura Sinica. 2015, 48(10):  1900-1914.  doi:10.3864/j.issn.0578-1752.2015.10.004
    Abstract ( 950 )   HTML ( 23 )   PDF (486KB) ( 1487 )   Save
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    Mapping crop patterns with remote sensing is of great implications for agricultural production, food security and agricultural sustainability. In this paper, the theoretical basis behind the mapping was summarized, mapping methods were classified into several categories, characteristics and applicabilities of different mapping methods in the latest decade were discussed intensively, and some important directions and priorities for future studies were proposed. Currently, spectral, temporal and spatial features are the major theoretical bases for crop pattern mapping. The mapping method based on single imagery is characterized by its simple implementation, but with difficulty of capturing imagery at the best time for distinguishing different crops. Instead, the mapping method based on time-series of imagery can make full use of temporal features and is thus widely used for crop mapping, among which the methods using multiple features are more suitable than the ones using a single feature for regions with complicated planting structure. To some extent, feature-oriented statistical modeling method can resolve the mixed-pixel problem but its robustness needs to be improved. Furthermore, large-scale crop pattern mapping can be done by combining the remote sensing and agriculture statistics. However, due to coarse resolution, the derived maps show poor region suitability. Future crop pattern mapping should target at developing “a map of crops”, the emphasis must be put on covering more crop types, enlarging the mapping areas, utilizing the superiority of blending multi-source data, strengthening the data preprocessing, optimizing the feature extraction and classifier selection, and improving the temporal and spatial scales of crop pattern mapping so as to better meet the needs of multi-faceted agricultural applications.
    Changes of Cropping System in China Based on Remotely Sensed NDVI data
    YANG Ting, ZHAO Wen-li, WANG Zhe-yi, LU Xing-tong, LU Shan
    Scientia Agricultura Sinica. 2015, 48(10):  1915-1925.  doi:10.3864/j.issn.0578-1752.2015.10.005
    Abstract ( 382 )   HTML ( 8 )   PDF (3685KB) ( 937 )   Save
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    【Objective】 The objective of this paper was to map the cropping systems of China using ten day NOAA- AVHRR- NDVI data of 1986 and 1996 years, and SPOT-VEGETATION-NDVI data of 2000. In order to extract the cropping systems information, the accuracies of the twice difference algorithm and Fourier transform method were compared. The performed better method was applied to map the cropping systems distributions for each year. On the basis of the cropping systems distributions, the changes of cropping systems in recent years would be known. Some meteorological data were used to reveal the possible reason of the change of cropping systems. The results were expected to provide information about the crop yield changes and their reasons.【Method】First, the cropping area was subset from the global land cover map provided by the European Space Agency (ESA GlobCover). Second, the cropping system distribution of 1986 was mapped using twice difference algorithm and Fourier transform method by ENVI and ArcGIS software. The accuracies of the two methods were compared using the cropping system information in the “Vegetation Map of China” at the scale of 1:4 000 000 as a reference. The cropping systems distributions of 1996 and 2006 were mapped by the twice difference algorithm. The distributions were compared to extract the change information of cropping systems. At last, the accumulated temperature indicators of the zero-grade zone were used to analyze the possible impact of the climate change on the cropping systems change.【Result】The twice difference algorithm and the Fourier transform both provided reasonable accuracy for mapping cropping system of China. The area of one crop a year was mainly in the northeast area, the two crops a year was distributed in Henan, Hebei, Shandong, and Anhui provinces, and the three crops a year was covered by the area south to the Hubei Province. The accuracy of twice difference algorithm (overall accuracy was 76.5% and Kappa coefficient was 0.64) was a little higher than the Fourier transform (overall accuracy was 69.4% and Kappa coefficient was 0.51) in this research. The northern limit of the two crops a year moved from Huludao, Liaoning Province in 1986 to Benxi and Shenyang, Liaoning Province in 2006. The western limit of it also moved from Pingliang, Gansu Province, Baoji, Shanxi Province in 1986 to Linxia, Ningxia Hui Autonomous Region and Haidong, Qinghai Province in 2006. The cross transformation was so complicated that the area of the one crop a year decreased by 330 thousand square kilometers, two crops a year increased by 360 thousand square kilometers, and three crops a year increased by 40 thousand square kilometers. The contour line of the accumulated temperature of the 4 200ºC and 6 100 ºC also moved to north and west in the studied years. The results may imply that the temperature is a main impact factor of changes of cropping systems.【Conclusion】Remote sensing data based on time sequence could be used to monitor real change of cropping system in China. The twice difference algorithm is a valid method to extract the crop systems information in China. The limit of the cropping system, especially for the two crops a year moved northwards and westwards. This movements resulted in the large scale decrease of one crop a year and significant increase of two crop a year, while the three crops a year did not change much. The global warming had an effect on the cropping system changes in China. Analyzing the distribution and change of cropping system in China by remote sensing data can not only help us understand and evaluate the crop production and production potential timely and accurately, but also benefit the agriculture-related departments for drawing up effective agricultural policies.
    The Possible Effects of Global Warming on Cropping Systems in China Ⅺ. The Variation of Potential Light-temperature Suitable Cultivation Zone of Winter Wheat in China Under Climate Change
    SUN Shuang, YANG Xiao-guang, ZHAO Jin, CHEN Fu
    Scientia Agricultura Sinica. 2015, 48(10):  1926-1941.  doi:10.3864/j.issn.0578-1752.2015.10.006
    Abstract ( 460 )   HTML ( 8 )   PDF (2388KB) ( 734 )   Save
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    【Objective】 In the context of climate change, China needs to address critical challenges relating to the agricultural development and food security. Wheat is one of China's three major grain crops, which plays an important role in China’s food security. The variation of potential light-temperature suitable cultivation zone of winter wheat in China under climate change was studied, which would provide an important scientific reference for the rational distribution of winter wheat. 【Method】 In this research, the ‘correction step by step’ method was used to calculate the potential light-temperature yield of winter wheat in each year. The year 1981 was taken as a time node and divided the period 1961-2010 into two sub-periods. Combined with the indices of the potential light-temperature yield level and the stability, the variation of the potential light-temperature yield level and the stability zones were investigated. In summary, the variation of the boundaries and distributions of the potential light-temperature suitable cultivation zones of winter wheat in China under climate change was studied. 【Result】 Compared with the period 1961-1980, during 1981-2010, the percentage of the very high and high potential yield zones in the study area increased by 6.33% and 7.42%, respectively, but the percentage of moderately high and marginally high potential yield zones decreased by 10.50% and 3.24%, respectively. During 1981 to 2010, the percentage of the very stable potential yield zone in the study area decreased by 25.76%, but the percentage of stable, moderately stable and marginally potential yield zones increased by 12.09%, 10.34% and 3.31%, respectively. The boundary of the very suitable zones moved northward and westward. The southern boundary of the very suitable zones moved northward from the central part of Anhui Province to the northwest portion of Shandong Province, and the western boundary of the very suitable zones extended westward from the southwest part of Shanxi Province to the northwest part of Shaanxi Province; The boundary of the suitable zones moved northeastward, and the maximum spatial displacement was in the eastern part of Sichuang Province to the southwest part of Shaanxi Province, moving by about 835.63 km. The spatial displacement of the potential suitable cultivation zone of winter wheat caused the changes of the winter wheat actual cropping area. Compared with the period 1961-1980, during 1981-2010, the percentage of the very suitable and suitable zones of winter wheat in the study area decreased by 4.49% and 4.75%, respectively, while the percentage of moderately suitable and marginally suitable zones increased by 7.17% and 2.06%, respectively. The potential yield per unit area increased by about 513 kg·hm-2 in average in each suitable cultivation zone. However, the total potential yield reduced by 1 504×104 t in the study area compared with 1961-1980 because of the reduction of the area in the very suitable and suitable zones of winter wheat, which caused the total potential yield in these two cultivation zones reduced by 5 108×104 t and 5 896×104 t, respectively. 【Conclusion】 Under the background of climate change, the percentage of the very high and high potential light-temperature yield zones of winter wheat increased, while the very stable potential light-temperature yield zone decreased. The very suitable and suitable zones of winter wheat in the study area decreased because of the decrease of stable; the boundary of the very suitable zone moved northward and westward, and the boundary of the suitable zone moved northeastward. The potential light-temperature yield per unit area increased because of the spatial displacement of the potential light-temperature suitable cultivation zone, which was beneficial to the improvement of the winter wheat yield. However, due to the reduction of the area in the very suitable and suitable zones of winter wheat, the total potential yield of winter wheat in China reduced. In the regions where the suitability of potential light-temperature yield has changed, the wheat production should be combined with the irrigation to ensure the high and stable yield.
    Agronomic Traits Variation Analysis of Huanghuai Dryland Winter Wheat under Temperature Change Background in China ——Taking Linfen, Shanxi as an Example
    LIU Xin-yue, PEI Lei, WEI Yun-zong, ZHANG Zheng-bin, GAO Hui-ming, XU Ping
    Scientia Agricultura Sinica. 2015, 48(10):  1942-1954.  doi:10.3864/j.issn.0578-1752.2015.10.007
    Abstract ( 556 )   HTML ( 16 )   PDF (507KB) ( 16538 )   Save
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    【Objective】Crop variety breeding was result from nature and artificial selection under climate change background. Huanghuai wheat region is the largest wheat region in China, that plays an important role for ensure wheat production and food security in China. The changes of agronomic traits of winter wheat and temperature in Huanghuai dryland in the past 30 years were studied in order to provide a theoretical basis and technological support for dryland wheat breeding to adapt to climate changes in the future. 【Method】 The agronomic traits data of control varieties and temperature data of Linfen in Shanxi province in the past 30 years were analyzed. Linfen is the national representative Huanghuai dryland winter wheat regional test site. The regularity of the main temperature traits of Linfen city, such as the average temperature and over 0 accumulated temperature in winter wheat growth, and agronomic traits of control varieties change trends were analyzed. The relationship between agronomic traits and meteorological elements were also analyzed by correlation, multiple regression and path analysis. 【Result】 The average temperature,over 0 accumulated temperature,the highest and lowest temperature was increased slowly year by year in wheat growth period. In wheat growth period, the average temperature was increased average 0.05 per year, over 0 accumulated temperature was increased average 21.9per year,there was positive relationship between the average temperature and over 0 accumulated temperature. Over 0 accumulated temperature change can reflect temperature change in whole wheat growth period. The highest and lowest temperature was increased 0.02and 0.16per year respectively. The average temperature and over 0 accumulated temperature was increased markedly from sowing to before the winter, the average temperature and over 0 accumulated temperature was increased significantly at vegetative growth stage, but were increased lightly and had a downward trend at reproductive stage. With climate warming, strong winter variety with more tillers was replaced gradually by winter and weak winter variety with moderate tiller. Agronomic traits of control variety were changed not much from 1986 to 1996, and changed greatly from 1997 to 2007, and changed markedly from 2007 to 2014, this change trend consist with temperature change trend in wheat growth period. In evolution of agronomic traits of Huanghuai dryland control varieties, the thousand-kernel-weight and kernels per spike was increased average 1.57% and 3.39% per year respectively, but the effective ear numbers and plant height as well grain yield were decreased average slowly 0.16% and 1.29% per year respectively. There was a significant positive correlation between grain yield and plant height (0.684**), effective ear number (0.531**) and thousand-seed-weight (0.541**). There were 46.73%, 26.17% and 3.26% variation of grain yield was decided by three main factors such as plant height and over 0 accumulated temperature as well as the average temperature from standing to jointing stage respectively. In Huanghuai dryland wheat high-yield breeding evolution, the over 0 accumulated temperature from standing to jointing stage and plant height had a higher positive effect on grain yield, but the average temperature from standing to jointing stage had a negative effect on grain yield. 【Conclusion】 The climate warming had a strong impact on agronomic traits evolution in Huanghuai dryland winter wheat region, selecting new wheat varieties with agronomic traits of middle plant height, middle effective ears numbers, more kernels per spike, and high grain weight is the improvement direction of middle-high yield and drought-resistant varieties in Huanghuai dryland wheat region for adapting to climate change in the future.
    PLANT PROTECTION
    Binding Characterization of Chemosensory Protein CSP1 in the Bemisia tabaci Biotype Q with Plant Volatiles
    WU Fan, ZHANG Xiao-man, ZHAO Lei, CUI Xu-hong, LI Hong-liang, LUO Chen
    Scientia Agricultura Sinica. 2015, 48(10):  1955-1961.  doi:10.3864/j.issn.0578-1752.2015.10.008
    Abstract ( 427 )   HTML ( 4 )   PDF (550KB) ( 704 )   Save
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    【Objective】The objective of this study is to clone the ORF (open reading frame) of chemosensory protein 1 (CSP1) from Bemisia tabaci biotype Q, and characterize the binding profiles of CSP1 with some candidate plants volatiles.【Method】 By means of full-length ORF primer, reverse transcription-polymerase chain reaction (RT-PCR) was used to amplify the full-length ORF BtCSP1, which was then cloned into the pET-30a (+)/BL21 (DE3) prokaryotic expression vector after double enzyme digestion. The recombinant BtCSP1 protein was expressed and purified by the method of Ni2+-agroase affinity chromatograph. The protein concentration was measured with Bradford method. The competitive fluorescence assay was used to analyze the binding properties of BtCSP1 with general plant volatiles with different chemical structures. As a suitable fluorescence reporter in studies of insect GOBPs’ function in vivo, N-phenyl-1-naphthylamine (1-NPN) was used to titrate the BtCSP1 solution until the fluorescence emission peak at maximum wavelength of BtCSP1 completely quenched. Then all plant volatiles were added into BtCSP1-1-NPN complex, respectively. The dissociation constants KD data represented the affinity of BtCSP1 with ligands were calculated by the Scatchard equation. 【Result】 BtCSP1 protein was successfully expressed after induction of 1 mmol·L-1 of IPTG, then purified by Ni2+ affinity column with gradient imidazole as washing solutions, finally dialyzed sufficiently using PBS buffer. The working concentration of BtCSP1 was diluted to 1.5 µmol?L-1. In the competitive fluorescence assay, the dissociation constants K1-NPNand the number of binding sites n of BtCSP1 and 1-NPN were 2.78 μmol·L-1and 0.82, respectively. It indicated that the binding of BtCSP1 with 1-NPN is strong and the binding ratio is almost 1:1. In all candidate chemicals, some plant volatiles preferred to bind with BtCSP1 with high affinity, such as 3-carene, 4-cymene, HL-cis-3-hexen-1-ol and α-pinene, which are reported to be related to the repellent characteristics of B. tabaci. Their binding affinity with BtCSP1 was strong, with the dissociation constant KD of 26.47, 39.43, 54.01 and 83.46 μmol·L-1, respectively. Especially 3-carene reduced the relative fluorescence intensity of 1-NPN by about 40% at the concentration of 200 μmol·L-1. 【Conclusion】 BtCSP1 exhibited a strong binding affinity with some repellent plant volatiles, indicating that BtCSP1 may be involved in the recognition of the repellent plants of host selection in the process of expanding into the invasion fields.
    The Alleviate Effect of Extracellular DNA and Protein in Maize Root Border Cells on the Allelochemical Stress from Chenopodium ambrosioides L.
    HU Zhong-liang, WANG Ya-nan, MA Dan-wei, CHEN Bin, HE Ya-qiang, ZHOU Jian
    Scientia Agricultura Sinica. 2015, 48(10):  1962-1970.  doi:10.3864/j.issn.0578-1752.2015.10.009
    Abstract ( 407 )   HTML ( 2 )   PDF (1105KB) ( 653 )   Save
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    【Objective】Root border cells (RBCs) are released from the root cap as individual cells or a group of attached cells. The mucilage of root border cells acts in a manner similar to that of neutrophil extracellular traps (NETs) in defense, thus, known as border cell extracellular traps (BETs). The extracellular DNA (exDNA) and proteins are components of border cell mucilage, which have been considered to play a vital role in protecting root tip from biotic and abiotic stresses. Allelopathy is one of the successful mechanisms of exotic plant for invading. The objective of this study is to reveal the roles of the exDNA and extracellular proteins of root border cell mucilage in resistance to allelochemical stress from an invasive plant, Chenopodium ambrosioides L..【Method】 Experiments were performed with maize (Zea mays L.) ‘Yayu26#’, a widely grown crop in the introduced habitats of C. ambrosioides under aeroponic culture with agar medium. The developmental characteristics of root border cells in maize were investigated. And the roots were treated with volatile oil from C. ambrosioides and its two main components, α-terpinene and cymene, respectively, then the experiments were divided into two groups. In one group, the root border cell viabilities and relative areas of root border cell mucilage were measured directly. In another group, the exDNA or extracellular proteins of root border cell mucilage were degraded by DNase I or protease, and then the cell viabilities and relative areas of root border cell mucilage of maize were studied.【Result】 The first border cell formation almost synchronized with root tip emergence in maize. The number of border cells increased with the development and elongation of the root. When the root length was about 30 mm, the number of border cells reached the maximum, that was about 6 130, then declined slightly to a stable level. Meanwhile, the activities of pectin methylesterase were correlated with root length negatively (P<0.01). Once the emerging of root, the activities of pectin methylesterase reached the top, and then declined into a low level. The cytotoxicity from volatile oil, α-terpinene and cymene resulted in significant decrease of the viabilities of root border cells (P<0.05). The allelopathic effects of α-terpinene and cymene were weaker than that of volatile oil. The allelopathic effects of mixed treatment group were different from the individual groups, which the viabilities of root border cells were higher than any other treatment groups, showed minimal toxicity. Volatile oil from C. ambrosioides had potential to induce the enlargement of mucilage with dose-dependent. At the dose of 5 μL, the relative areas of mucilage enlarged significantly in volatile oil treatment groups (P<0.05). Compared with the control, α-terpinene-treated and cymene-treated induced no significant change of the relative areas of mucilage. Degradation of exDNA or extracellular proteins with DNase I or protease resulted in loss of root border cells resistance to allelochemical stress, and the viability of root border cells decreased obviously, there was less than 10% viabilities of root border cells remained in volatile oil treatment groups, while there was more than 80% in α-terpinene-treated, cymene-treated and the combined treatment of α-terpinene and cymene. 【Conclusion】Volatile oil from C. ambrosioides and its two main components, α-terpinene and cymene were cytotoxical, both of them inhibited viabilities of root border cells, while exDNA and extracelluar proteins of root border cell mucilage play an important role in resistance to allelochemical stress from C. ambrosioides.
     
    Molecular Cloning and Expression Profiling of a Ryanodine Receptor Gene in the Peach Fruit Moth (Carposina sasakii)
    SUN Li-na, ZHANG Huai-jiang, YAN Wen-tao, MA Chun-sen, QIU Gui-sheng
    Scientia Agricultura Sinica. 2015, 48(10):  1971-1981.  doi:10.3864/j.issn.0578-1752.2015.10.010
    Abstract ( 416 )   HTML ( 2 )   PDF (5940KB) ( 493 )   Save
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    【Objective】Ryanodine receptor (RyR), the target of diamide insecticides, is the largest known iron channel protein. The receptor is the key to the stabilization of Ca2+ by regulating the release of Ca2+ in cell. The objectives of this study are to isolate the cDNA of ryanodine receptor from the peach fruit moth Carponsina sasakii (CsRyR), and to analyze the expression of RyR mRNA at different developmental stages of this pest. 【Method】 According to the sequences of RyR gene of other species obtained from the NCBI database, degenerate and specific primers were designed for reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends PCR to clone the full-length sequences of CsRyR. The open reading frame (ORF), animo acid residues, the conserved structure domains, phylogenetic tree and other characteristics were analyzed using the bio-software. And the relative expression levels of RyR mRNA at different developmental stages (egg, larvae, pupe and adult) were investigated using quantitative real-time PCR (RT-qPCR), GAPDH was used as the reference. 【Result】 The full-length cDNA sequence of CsRyR was isolated from the peach fruit moth using RT-PCR and RACE-PCR. The CsRyR mRNA has an open reading frame (ORF) of 15 405 bp nucleotides and encodes 5 134 amino acid residues. CsRyR displays 45%-47% identity with vertebrate RyR isoforms, and 46% identity with RyR from Caenorhabditis elegans (CeRyR). And CsRyR shares 91%-94% identity with that of Lepidoptera insects and 79% identity with those of Homoptera and Diptera insects. Meanwhile, the phylogenetic tree analysis showed that CsRyR has the closest genetic relationship with insects of Noctuidae and Pyralididae.CsRyR shares common structural features with known RyRs. Six transmembrane domains (AA 4 467-5 029) are at the COOH-terminal. The sequence motif, GVRAGGGIGD, which constitutes part of the pore-forming segments of the Ca2+ release channels. The sites of RyR binding with Ca2+ and ATP (EF-hand and GXGXXG (3 motifes)) exist. Furthermore, three putative sites potentially binding with diamides were AA 183-290 (BmRyR), AA 4 610-4 655 (DmRyR) and 4 946 G (PxRyR) showed no differences in the CsRyR. And the relative expression abundance of RyR mRNA from the pupae of C. sasakii was the highest, which were 25.19, 7.73, 6.48, 4.74 and 3.58 fold compared to 1-day-old eggs, 6-day-old eggs, infant larvae, mature larvae and adults, respectively. 【Conclusion】 The full length of RyR cDNA was cloned from C. sasakii, and the mRNA expression has significant differences at different developmental stages.
    SOIL & FERTILIZER·WATER-SAVING IRRIGATION·AGROECOLOGY & ENVIRONMENT
    Soil Moisture and Enrichment Regularity of Steppe Soil in Qinghai Lake Area
    MA Yan-dong, ZHAO Jing-bo, SHAO Tian-jie, XING Shan
    Scientia Agricultura Sinica. 2015, 48(10):  1982-1995.  doi:10.3864/j.issn.0578-1752.2015.10.011
    Abstract ( 505 )   HTML ( 1 )   PDF (1239KB) ( 406 )   Save
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    【Objective】The aim of this study on the steppe soil in Qinghai Lake area is to explore the soil moisture migration and its enrichment characteristics, soil moisture distribution model, hydrological cycle and balance and reveal the characteristics of soil reservoirs, dried soil layer and its recovery conditions, and then provide a scientific basis for protecting soil water resources and steppe vegetation, construction of soil reservoir and sustainable development of grassland ecological environment.【Method】A series of experiments were performed to determine soil moisture, soil suction, infiltration rate and particle size. More than 600 soil moisture samples were collected by Light-duty Human Drill in 2009-2012. Drying weighing method was used to determine soil water content. Bicyclic infiltration method was used to determine soil infiltration rate in situ. Soil particle size was analyzed by Laser Particle Analyzer. Soil suction was measured by Tensiometers in situ.【Result】The distribution of soil water at soil profile has a certain stability, that is, in dry season or rainy season, the soil water in this area is enriched about 65% at 0-0.4 m depth and is quite shortage under 0.6 m depth. The soil suction in this area ranging from 0.17 MPa to 0.42 MPa suggests that field capacity of this area is about 20%. The infiltration rate of soil in this area varying from 1.3 mm·min-1 to 3.0 mm·min-1 shows that it is high and advantageous to precipitation into soil. The soil moisture is about 23% and 6.5% at 0-0.4 m depth and under 0.6 m depth, respectively. Field capacity of this area is about 20%, meaning that there is about 3% gravity water at 0-0.4 m depth of soil. The soil usually develops a dried soil layer with different levels under about 0.6 m in this area, and the greater soil thickness, the more seriously dried soil layer develops. Under 0.4 m depth of soil, the relationships between soil moisture and depth can be described by power function. The parameters of power functions (i.e. a, k) are able to well reflect the average level of soil moisture (a) and the degree of soil moisture decreased with the increase of depth (k). As the precipitation increased by about 50 mm from 2009 to 2011, the increment of soil moisture can be clearly reflected by the incremental curve of analogue function. The increment of soil moisture within the depth from 0.4 m to 0.8 m gradually decreased from 5% to 3% and was less than 3% under 0.8 m depth. The soil property is excellent in this area, but the soil moisture under 0.6m depth is close to or below 5% unavailable water of silty soil.【Conclusion】The retention and enrichment of soil water at 0-0.4 m depth is rare and this characteristic mainly depend on the low temperatures and long-term freezing of soil in Qinghai Lake area. The regulation function of soil reservoir is weak in this area due to the average thickness of soil is less than 1.5 m. To some extent, the retention and enrichment of soil water not only can enhance the regulation function of soil reservoir for vegetation in shallow root, but also can play an important role in inhibiting the occurrence of grassland desertification. The soil water balance in this area showed a weakly positive balance because of 400-420 mm annual precipitation from 2009 to 2011. Under this condition, the dried soil layer in thin soil was able to disappear completely, but could not in thick soil. The dried soil layer was recovered slowly in this area and its soil moisture recovered was less than 5.0%. Development of dried soil layer and its distribution with small depth not only show that the ecosystem in this area is more vulnerable, but also indicate this area is not suitable for growing arbor vegetation because it will consume more water.
    Effects of Water and Nitrogen Coupling on Root Growth and Single Fruit Weight of Greenhouse Muskmelon
    YUE Wen-jun, ZHANG Fu-cang, LI Zhi-jun, WU Li-feng
    Scientia Agricultura Sinica. 2015, 48(10):  1996-2006.  doi:10.3864/j.issn.0578-1752.2015.10.012
    Abstract ( 464 )   HTML ( 3 )   PDF (489KB) ( 9007 )   Save
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    【Objective】The objective of this experiment is to study the effects of different water and nitrogen levels on root growth and a fruit weight of greenhouse muskmelon, to explore the relationships between root growth, single fruit weight and water and nitrogen supply, and to provide a scientific basis for water and reasonable nitrogen application of local greenhouse muskmelon.【Method】According to the environment parameters (light, temperature, humidity, etc.) inside the solar greenhouse, ‘Yi pin tian xia 208’ of muskmelon was chosen as the test cultivar, irrigation application amount was determined by using modified Penman- Monteith equation. A total of 9 treatments were designed. Muskmelon plants were subjected to three irrigation treatments, that wass 70%, 100% and 130% of crop ETc (Evapotranspiration) and three N levels, including low nitrogen level N1 (70 kg N·hm-2), traditional nitrogen level N2 (130 kg N·hm-2) and high nitrogen level N3 (180 kg N·hm-2). Treatments were completely arranged randomly. A greenhouse muskmelon crop was grown under mulched drip irrigation conditions to investigate the effects of different water and nitrogen levels on root growth, distribution and single fruit weight.【Result】 Most of the total root lengths were concentrated on the 0-30 cm soil layer. Total root length increased slowly as soil layer increased. Total root length, single fruit weight and water use efficiency of muskmelon descended with the rising of irrigation water amount at the same nitrogen level. The highest values of total root length and single fruit weight were observed in W2N2, which were 6 625.48 cm and 818.94 g, respectively. Total root length and single fruit weight of muskmelon showed a trend of decreasing after the first increasing with the rising of irrigation water amount at the same nitrogen level. Water use efficiency descended with the rise of irrigation application rate, PFPN descended with the rise of nitrogen application rate. There was a significant relationship between root length with less than 2 mm, root dry weight and yield,and the correlation reached a significant level, the more root growth, the higher yield would increase, reasonable amount of irrigation water and nitrogen fertilizer could promote root system to absorb water and nutrients, thus increasing production.【Conclusion】The change rule of total root length in the depth of root vertical direction could be simulated by the equation y=A(1-Bx), the equation determination coefficient R2 reached more than 0.9. The principle component analysis method was used to evaluate the root growth of muskmelon, and the results showed that comprehensive principle component could represent 97.27% total root information, the highest comprehensive evaluation was obtained in W2N2. Unreasonable irrigation and nitrogen applications could result in reducing single fruit weight, characteristic parameters of muskmelon root, water use efficiency and partial factor productivity from applied N. It was concluded that the irrigation level W2 and nitrogen level N2 could be recommended as the best combination of water and nitrogen, which promoted the root growth, improved single fruit weight, water and nitrogen use efficiency of muskmelon in the muskmelon production under drip irrigation with plastic film mulching in the experimental area.

    HORTICULTURE
    Analysis of Expression Levels of Floral Genes in the Buds on Different Branch Nodes of Grapevine
    LIU Dan, SUN Xin, MU Qian, WU Wei-min, ZHANG Zhen, FANG Jing-gui
    Scientia Agricultura Sinica. 2015, 48(10):  2007-2016.  doi:10.3864/j.issn.0578-1752.2015.10.013
    Abstract ( 352 )   HTML ( 2 )   PDF (760KB) ( 657 )   Save
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    【Objective】 The objective of the paper is to study the expression pattern of eight genes in different nodes and the phenological period of buds development, and explore the mechanism of the time and speed of buds development on different nodes of grape branches.【Method】 In this study, the expression of eight floral genes (VvFT, VvSOC1, VvAP1, VvAP2, VvAP3, VvFUL, VvAG and VvFLC) in the buds at different branch nodes of 8-year-old ‘Fujiminori’ grapevine was examined by real-time quantitative PCR, and the phenological period of flower bud differentiation was observed. 【Result】The floral differentiation of ‘Fujiminori’ grapevine showed a remarkable characteristics of ‘beyond section’. The first bud began to form and develop from late April or early May, and the formation time of the last bud at the top was in late September. The development time of the lower nodes was five months longer than the upper nodes, but the flowering time of them was almost at the same in the second season. The buds in the upper position differentiated from the bottom upwards to the top gradually, and the process of bud differentiation at the top was shorter than that at the bottom. The expression of floral genes in buds at different branch nodes was different. The expression level of VvFT was low and showed no significant difference during the growing season. VvSOC1 , VvAP2, VvAP3 and VvFUL. The highest expression level appeared in the early period of bud differentiation, and then decreased over time. Buds on middle nodes undergo a low speed differentiation process for a long time, and the expression of floral genes kept a high level during the differentiation period. By contrast, the buds on upper and lower nodes differentiated at a higher speed, and the expression of floral genes was lower. In the end, the development of the buds on different nodes reached the similar level. 【Conclusion】 The expression levels are different on different nodes, and high expression periods were also different on different nodes. Although the peak period of floral development of lower and upper buds were relatively late, continuous development was observed in these nodes, which led to the floral differentiation simultaneously on the lower, middle and upper nodes. This may be one of the factors that lead to difference of differentiation quality between buds on different nodes.gene in the buds at different branch nodes expressed highly during all the growing season, and showed a similar expression trend. The expression of VvAP1, VvAP2, VvAP3, and VvFUL genes in the buds at lower and upper nodes showed no remarkable change, however on the middle nodes, the expression of these genes changed greatly, and appeared an obvious upgrade firstly then descending latter tendency, what’s more, in the buds on 8, 11 and 15th nodes, the expression of these genes was higher than that at other nodes. VvAG showed different expression patterns with VvAP1
    The Core Collection Construction of Xinjiang Wild Apricot Based on ISSR Molecular Markers
    LIU Juan, LIAO Kang, ZHAO Shi-rong, CAO Qian, SUN Qi, LIU Huan
    Scientia Agricultura Sinica. 2015, 48(10):  2017-2028.  doi:10.3864/j.issn.0578-1752.2015.10.014
    Abstract ( 433 )   HTML ( 1 )   PDF (553KB) ( 604 )   Save
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    【Objective By comparison analysis of the results of different sampling strategies and genetic distances, the method for constructing core collection of Xinjiang wild apricot based on molecular markers data was studied in order to establish the optimum core collection resources which will be beneficial to the protection and use of resources. 【Method】 Taking 135 wild apricots as materials that were come from Huocheng Daxigou, Xinyuan Boersai and Gongliu Yiligedai in Xinjiang, using UPGMA stepwise clustering method according to SM, Jaccard and Nei&Li genetic distances for initial collection, until any one sampling location was no collection into the core collection, then an allele preferred sampling strategy was used to construct wild apricot core collection and compared with the random sampling strategy. The number of lost allele and t-test of number of polymorphic loci, percentage of polymorphic loci, observed number of alleles, effective number of alleles, Nei’s genetic diversity and Shannon information index for gene diversity were used to determine the optimal building methods. T-tests of core collection, initial collection and reserve collection were conducted, and the genetic diversity of them was compared to evaluate the representative of core collections. The principal coordinate analysis method and the phenotypic traits of initial and core collections were used to confirm the core germplasm.【Result】Compared with the random sampling strategy, allele preferred sampling strategy could construct more representative core collections that with higher values of genetic diversity indexes and little polymorphic loci. According to Nei&Li genetic distance, the constructed core collection of Xinjiang wild apricot was better than by SM and Jaccard genetic distance for these core collections had high values of genetic diversity indexes. The analysis of principal coordinate analysis and phenotypic traits showed that the core collection of Xinjiang wild apricot constructed by allele preferred sampling strategy and Nei&Li genetic distance could more comprehensively represent at the genetic diversity of wild apricot initial collection. The 31 core collection resources of wild apricot includes 22.96% germplasm samples of the initial collection, the retention ratio of number of polymorphic loci, percentage of polymorphic loci, observed number of alleles, effective number of alleles, Nei's genetic diversity and Shannon information index were 92.69%, 98.83%, 99.42%, 103.26%, 109.24% and 108.31%, respectively. 【Conclusion】 The method of allele preferred sampling strategy and Nei&Li genetic distance by stepwise clustering is a suitable method for constructing Xinjiang wild apricot core collection. These results demonstrated that the 31 core collection could stand for original collection excellently, at the same time this research method of the construction of core collection has important reference values for other crops.
    STORAGE·FRESH-KEEPING·PROCESSING
    Effects of Different Storage Methods on Postharvest Physiology and Storage Quality of Fresh Walnut Fruit
    YANG Xi, ZHANG Run-guang, HAN Jun-qi, ZHANG You-lin
    Scientia Agricultura Sinica. 2015, 48(10):  2029-2038.  doi:10.3864/j.issn.0578-1752.2015.10.015
    Abstract ( 580 )   HTML ( 4 )   PDF (1373KB) ( 804 )   Save
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    【Objective】 Fresh walnut fruit easily appears decay and browning, its kernel easily becomes mildewed and dehydrated during storage. The supply period of fresh walnut fruit is short and walnut used in production is often conserved by dry. The objective of this study is to provide a theoretical basis and technical method for prolonging the supply period of fresh walnut through studying its postharvest physiology and storage technology. 【Method】 ‘Xiang ling’ walnut variety was used as raw material in this study, air flow method was used to analyze the respiratory intensity of fresh walnut with husk and fresh walnut without husk. Moreover, the respiration type was also studied. Before storage under the condition of temperature (1.0±0.5)℃, RH 90%-95%, the materials were subjected to storage in vacuum-package with plastic bag, modified atmosphere package with plastic bag, controlled atmosphere storage of text box, respectively. A contrast was prepared without any treatment. To study the effect of gas composition on postharvest physiology and storage technology of fresh walnut fruit, moisture content, reducing sugar content, lipid content, protein content, MDA content and oil acid value, iodine value, peroxide value of fresh walnut kernel were measured during storage. Related enzymes of fresh walnut kernel such as LOX activity, SOD activity and CAT activity were determined, too. Moreover, green husk browning index of fresh walnut fruit was evaluated in different periods of storage. After storage for 120 d, walnut shell color, endotesta color, kernel flavour of total fresh walnut fruit were evaluated, respectively. Then, the appearance of fresh walnut husk, shell and kernel were taken photo. 【ResultThe results showed that the fresh walnut fruit with green husk is respiration climacteric fruit and fresh walnut without green husk is non-respiration climacteric fruit. The content of moisture, lipid, and oil iodine value presented a reduced trend, but the acid value, peroxide value, LOX activity, and MDA content of fresh walnut kernel showed a increased trend and SOD activity, CAT activity, reducing sugar content, protein content showed no visible change. The contrast treatment showed the worst preservation effect and green husk browning index reached 0.27 after storage for 40 d. However, the green husk browning index of vacuum-package with plastic bag storage was only 0.18 after storage for 120 d. The result also showed that the treatment of vacuum-package with plastic bag could effectively prevent browning of green husk, maintain the moisture of kernel, slow down the change rate of oil acid value, iodine value and peroxide value, keep the activities of SOD and CAT at a high level, inhibit the generation of MDA. By evaluating the sensory quality after storage for 120 d, the treatment of vacuum-package with plastic bag gained the highest score in every single project. 【Conclusion】 The lower the concentration of O2 in storage circumstances, the better the effect of conservation. After plucked with carpopodium in walnut commercial mature period, pre-cooled at 4.0℃ for 3 d, and vacuumed with LDPE plastic bag using vacuum pack machine, the storage life of walnut could reach 120 d at(1.0±0.5)℃ and RH 90%-95%. Its pericarp browning index of fresh walnut was low, and the color, aroma, taste and shape remained good.
    ANIMAL SCIENCE·VETERINARY SCIENCERE·SOURCE INSECT
    Differentially Expressed microRNAs Screening Between Ovaries of Sheep Producing Single Lamb and Twins
    QI Yun-xia, LIU Xiao-fang, ZHANG Ping, HE Xiao-long, XING Yu-mei, Dalai, Terigele, LIU Yong-bin, RONG Wei-heng
    Scientia Agricultura Sinica. 2015, 48(10):  2039-2048.  doi:10.3864/j.issn.0578-1752.2015.10.016
    Abstract ( 365 )   HTML ( 1 )   PDF (1959KB) ( 638 )   Save
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    【Objective】This study was conducted to get a comprehensive understanding of miRNAs expression in sheep ovary, and to analyze their expression differences between sheep producing single lamb and twins, thus providing a basis for exploring the role of miRNAs in the regulation of fertility. 【Method】 Multi-species miRNA microarray was used to profile ovarian miRNAs expression in sheep. Firstly, single-lamb-producing and twin-producing ewes were selected. Ovaries were collected after estrus for total RNA extraction, and small RNA fragments were isolated and hybridized with miRNA microarray. Then data analysis was performed to obtain sheep ovarian miRNAs profile. Differentially expressed microRNAs were selected by bioinformatics software with the standard of q-value ≤ 5% and Fold Change ≥2 or ≤0.5, and q-PCR technique was employed to testify the microarray results. Two different methods, microT and miRDB, were used, respectively, and then data were merged and to predict target genes. Finally, online software was used to perform GO function annotation for target genes. 【Result】 Among all the detected miRNAs, a total of 5448 miRNAs were detected in sheep ovaries, of which 22 were specifically expressed in single-lamb-producing group, while 15 were in twin-producing group; For the miRNAs reported in sheep, expression differences were compared between two groups, and obtained 11 differentially expressed miRNAs, with 7 down-regulated and 4 up-regulated. Tow miRNAs were randomly selected to perform q-PCR validation, and the results were consistent with the microarray results, illustrating that the microarray results were accurate and credible. Target genes were obtained for 7 miRNAs: oar-miR-370-5p, oar-miR-376b-5p, oar-miR-381-5p, oar-miR-412-5p, oar-miR-541-3p, oar-miR-544-5p and oar-miR-1185-5p,and the number of target genes for each miRNAs was: 115, 71, 1, 5, 8, 135 and 23. The GO annotation results showed that target genes of miR-376b-5p and miR-1185-5p are mainly involved in forming intracellular components and organelles; in the molecular function category, the vast majorities of genes are the connection and catalytic activity molecules; in biological process, mainly involved in cell proliferation, differentiation, apoptosis and metabolism. Meanwhile, the target genes of miR-376b-5p ( IGF-1, DAZL, MTOR, MET and NEDD4 ) and miR-376b-5p ( AHR ) take part in reproduction regulation.【Conclusion】Sheep ovarian miRNAs expression profiling was successfully constructed, and differentially expressed miRNAs between single-lamb-producing and twin-producing sheep were obtained. These miRNAs may be involved in sheep follicular development and litter size regulation.
    Effect of Dietary Protein Sources on Egg Performance and Egg Quality of Jinghong Laying Hens at Peak Production
    WANG Xiao-cui, ZHANG Hai-jun, WU Shu-geng, YUE Hong-yuan, LI Jie, QI Guang-hai
    Scientia Agricultura Sinica. 2015, 48(10):  2049-2057.  doi:10.3864/j.issn.0578-1752.2015.10.017
    Abstract ( 484 )   HTML ( 11 )   PDF (392KB) ( 1252 )   Save
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    【Objective】 This experiment was conducted to evaluate the effects of dietary protein sources on egg performance and egg quality of Jinghong laying hens at peak production. 【Method】 Two hundred and eighty-eight Jinghong laying hens aged 32 weeks with similar egg production were randomly divided into 4 groups with 6 replicates per group and 12 hens per replicate. Hens in group Ⅰ (control group) were fed soybean meal (SBM) protein, hens in group Ⅱ were fed degossypolized cottonseed protein (DCP), hens in group Ⅲ were fed double-zero rapeseed meal (DRM) protein, and hens in group Ⅳ were fed 4% yellow mealworms (YM) and soybean meal (SBM) protein. All diets were isocaloric (11.11 Mcal·kg-1), isonitrogenous (CP, 16.5%) and had the similar standard ileum digestible AA model. The experiment lasted for 8 weeks. 【Result】 (1) In the whole feeding period,no significant differences were found on average egg production, egg weight and cracked eggs ratio among all groups (P>0.05). In DCP group, egg mass during 1-8w was lower than that SBM group or 4% YM group (P<0.05), while the egg mass of DRM group was lower than that SBM group(P=0.084). The average feed intake of DCP group and 4% YM group during 1-8w were lower than that SBM group (P=0.091), while the average feed intake of DRM group was lower than that SBM group(P=0.091). Egg/feed also showed that the same tendency in all groups, that is the egg/feed of DCP group and the DRM group were reduced (P=0.052), while the egg/feed of 4% YM group had no significant difference (P>0.05). (2) No significant differences in the plasma aspartate transaminase (AST) among all groups were observed (P>0.05). Compared to SBM group, plasma alanine transaminase (ALT) in laying hens of 4% YM group was significantly reduced at 8w (P<0.05). The plasma total protein, globulin in laying hens of DCP group at 8w were significant lower than that SBM group (P<0.05). The plasma albumen in laying hens of 4% YM group was higher than those DCP group and DRM group at 4w and 8w (P<0.05). (3) There was a trend toward eggs of DCP group and DRM group at 8w decreasing shell strength (P=0.084) and shell thickness (P=0.062), while the egg of 4% YM group increasing shell strength (P=0.084) and shell thickness (P=0.062). Compared to SBM group, albumen height of eggs in DCP group and DRM group were reduced 4.02% and 3.41% (P=0.091) at 4w, while the 4% YM group was increased 3.87% (P=0.079). No significant difference were found albumen pH among all groups (P>0.05). (4) Albumen percent of eggs in DCP group and DRM group were reduced 1.67% and 1.18% (P=0.099) than that SBM group at 4w, while 4% yellow mealworms group was increased 1.70% (P=0.099). In addition, yolk dry matter of egg in 4% YM group was higher than that DRM group at 4w (P<0.05). 【Conclusion】 4% yellow mealworms replace soybean meal and soybean meal was the most optimal protein ingredients for Jinghong laying hens at peak production, followed by double-zero rapeseed meal and degossypolized cottonseed protein.
    Antimicrobial Resistance Investigation and Dominant Genotype Analysis of ESBLs- Producing Escherichia coli Strains from Chicken in Qingdao
    QU Zhi-na, LIU Hong-yu, WANG Juan, ZHAO Si-jun, LI Yu-qing, HUANG Xiu-mei, GAI Wen-yan, WANG Jun-wei
    Scientia Agricultura Sinica. 2015, 48(10):  2058-2066.  doi:10.3864/j.issn.0578-1752.2015.10.018
    Abstract ( 387 )   HTML ( 1 )   PDF (1424KB) ( 560 )   Save
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    【Objective】 By using antimicrobial susceptibility testing and detection of phenotype and genotype in extended spectrum beta-lactamases (ESBLs), this study aims to find out the distribution and drug-resistance of ESBLs-producing E.coli strains on broiler farms in Qingdao, and then analyze genotype and gene subtype of ESBLs, so as to provide a basis for guiding rational clinical use of antimicrobial agents and effective control of infection and spread of ESBLs-producing strains.【Method】The antimicrobial susceptibility of 249 E. coli strains was determined by micro broth dilution, the phenotype of ESBLs was identified and determined by CLSI standard method, DNA was amplified and the genotype of drug resistance plasmid of ESBLs was analyzed by PCR, sequencing and biological software, and the significant difference between ESBLs-producing and non-producing strains was analyzed  with SPSS19.0 software.【Result】There were 83.13% of E.coli strains produced ESBLs. The resistance rate of ESBLs-producing strains to 7 kinds of drugs was higher than that of non-ESBLs-producing ones, and there was a significant difference between them (P<0.05) to GM, SPT, AM, A/C and CEF. But the resistance rate of ESBLs-producing strains to tetracycline and florfenicol was significantly lower than those in non-ESBLs-producing ones. The multi-drug resistance rate of ESBLs producing and non-producing strains was 99.03% and 92.86%, respectively (P=0.035). The detection rate of CTX-M, TEM and OXA genotype was 100%, 99.52% and 47.83%, respectively, and SHV genotype wasn’t detected. The enzyme producing strains belong to 10 gene subtypes, the dominant ones of which were TEM-1, CTX-M-65, CTX-M-55, and OXA-1. CTX-M-123 and CTX-M-64, the recombinant chimeras, were found from healthy poultry for the first time. 【Conclusion】The chicken origin ESBLs-producing E.coli strains are widely spread and transmitted in Qingdao. The antimicrobial resistance of ESBLs-producing strains are more serious than that of non-ESBLs-producing strains. Compared to other areas in China, CTX-M and TEM are also popular genotypes in Qingdao, but with different gene subtypes, TEM-1, CTX-M-65, CTX-M-55 and OXA-1 are dominant gene subtypes of each genotype, respectively.
    RESEARCH NOTES
    Effect of Brassinolide on Calcium Ion Distribution of Plant Cell
    HUANG Xin, GAO Meng-zhu, ZHANG Hao, GAO Jing, WANG Feng-ru, DONG Jin-gao
    Scientia Agricultura Sinica. 2015, 48(10):  2067-2075.  doi:10.3864/j.issn.0578-1752.2015.10.019
    Abstract ( 345 )   HTML ( 1 )   PDF (2346KB) ( 883 )   Save
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    【Objective】The objective of this study is to analyze the effect of Brassinosteroid (BR) on Ca2+distribution of plant cell, analyze the influence of BR on the expression level of the genes coding Ca2+ channel and Ca2+-ATPase, and clarify the effect of BR on calcium homeostasis in Arabidopsis thaliana. 【Method】The effect of BR on Ca2+ location in plant cell was studied using the potassium pyruantimonatc technique. The expression level of genes which coded Ca2+-ATPase (located in the membrane, vacuole, endoplasmic reticulum) and Ca2+ channel (located in membrane, vacuole, lysosome) was studied by Real-time PCR technique.【Result】Under normal condition, Ca2+ mainly distributed in cell walls, intercellular space, vacuole, and there were only a few Ca2+ distributed in the chloroplast and cytoplasm in plant cells. After 1 µmol·L-1 BR treatment for 3 hours, Ca2+ gathered nearby vacuole membrane and cell membrane, meanwhile, the distribution of Ca2+ in the cytoplasm and chloroplast was increased. After BR treated for 6 hours, the Ca2+ distribution in the cytoplasm and chloroplast was continue increased, the Ca2+ distribution in the cell walls was reduced; after BR treated for 9 hours, the Ca2+ distribution in the cytoplasm and chloroplast was decreased, but increased in the intercellular space and vacuole, while the Ca2+ distribution in the cell walls was obviously reduced. It suggested that BR had function to release Ca2+ from cell wall. CNGC2 and CNGC12 are genes which encoded Ca2+ channel in the cell membrane. After 1 µmol·L-1 BR treated for 3 hours, the expression level of CNGC2 and CNGC12 were all obviously decreased, this indicated that BR could block the extracellular Ca2+ transfer into cytoplasm. After 1 µmol·L-1 BR treated for 6 hours, the expression levelsof CNGC2 and CNGC12 recovered;after 1 µmol·L-1 BR treated for 9 hours, the expression levels of CNGC2 and CNGC12 were obviously increased. TPC1 and TPC2 are genes encoding Ca2+ channel in the vacuole and lysosome, respectively. The expression levels of TPC1 and TPC2 were significantly decreased after 1 µmol·L-1 BR treated for 3 hours, but the TPC1 expression level was obviously higher than that without BR treated; the expression level of TPC2 was also significantly increased after 1 µmol·L-1 BR treated for 9 hours. The results showed that BR could block the fast rising of the cytoplasm Ca2+ Concentrations, the expression level of the genes coding vacuole Ca2+ channel recovered earlier than that of the Ca2+ channel genes in the cell membrane and lysosome. ACA8 and ACA10 are genes encoding Ca2+-ATPase located in the cell membrane. After 1 µmol·L-1 BR treated for 3 and 6 hours, the expression level of ACA8 and ACA10 had no significant change. After BR treated for 9 hours, the expression level of ACA8 and ACA10 was significantly increased. ACA4 and ACA11 were genes encoding Ca2+-ATPase located in the vacuole membrane, the change of ACA4 and ACA11 expression level was similar to the change of ACA8 and ACA10. ACA2 was one gene encoding Ca2+-ATPase located in the endoplasmic reticulum, the expression level of ACA2 also appeared the highest peak after 1 µmol·L-1 BR treated for 9 hours. The results indicated that the expression level of Ca2+-ATPase genes increased after BR treated for 9 hours, and high concentrations cytoplasm Ca2+ was infused into extracellular and intracellular calcium store including intercellular space, vacuole, endoplasmic reticulum, etc. So BR could regulate the cytoplasm calcium homeostasis.ConclusionThe second messenger Ca2+ was regulated by BR, and the signaling of BR was conducted through regulating the calcium homeostasis regulation system.
    Effect of Piperine- and Sanshool-Experiences on Larval Feeding of Helicoverpa armigera
    LI Wei-zheng, HU Jing-jing, LI Hui-ling, GUO Xian-ru, YAN Feng-ming, YUAN Guo-hui
    Scientia Agricultura Sinica. 2015, 48(10):  2076-2084.  doi:10.3864/j.issn.0578-1752.2015.10.020
    Abstract ( 315 )   HTML ( 2 )   PDF (463KB) ( 856 )   Save
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    【Objective】Feeding deterrents or anti-feedants are a broad range of chemicals regulating the feeding behaviour via altering the palatability of crops to herbivores. The idea of pest ecological control using feeding deterrents is an alternative strategy to overcome the negative effect of chemical control. However, almost all insect pests can develop experience-dependent adaptivity to various feeding deterrents, especially gustatory habituation, which limited the practical use of these substances. The objective of this study is to elucidate the sensory organs involved in the feeding deterrence of two amides against a generalist herbivore, Helicoverpa armigera (Hübner), and the effect of prior feeding experience of piperine and sanshool on subsequent feeding behaviour of the larvae, providing a scientific basis for large-scale application.【Method】The olfactory approaching response under dual-choice condition with piperine- and sanshool-treated with tobacco leaf discs was tested, respectively, paired with the control leaf disc, and the feeding duration of the larvae after first encounter with the piperine- or sanshool-treated tobacco leaf discs under non-choice condition was measured. Then the effect of larval feeding experience to piperine and sanshool on their subsequent choice feeding behaviour was tested based on the concentrations at 50% feeding deterrence against the fourth instar larvae. 【Result】Piperine and sanshool did not exhibit any repellent effect, and the feeding durations of the larvae after first encounter with piperine- and sanshool-treated tobacco leaf discs were approximately 30 s, significantly shorter than that of the control (longer than 100 s). This feeding duration time was in accordance with the typical post-ingestive response time of a model insect species, Manduca sexta. The concentrations at 50% feeding deterrence to the fourth instar larvae of piperine and sanshool were 0.2259 and 0.4003 mg per tobacco leaf disc (1.5 cm ID), respectively. The bioassay of feeding experience showed that the larvae with piperine- and sanshool-feeding experiences were significantly deterred by the corresponding experienced substances, suggesting that no significant habituation occurred when the two amides were mixed with the artificial diet at the concentrations of 50% feeding deterrence to the fourth instar larva and when larval feeding experience was induced from the third instar to the fifth instar. The larvae experienced with sanshool were also deterred significantly by piperine. However, the larvae experienced piperine developed gustatory habituation to sanshool, demonstrating an asymmetric cross-habituation between these two amides. The binary mixture of piperine and sanshool could not mitigate the gustatory habituation of the larvae, whether considering the difference between the feeding consumption of the treated and control tobacco leaf discs within each experience group, or considering the feeding deterrence rates between of different experience group when tested under the same conditions. The possible reasons were discussed with respect to the induced stage and the mechanism of the tested substances deterring feeding. The binary mixture of piperine and sanshool could not mitigate the decrease in feeding deterrent response following prolonged exposure.【Conclusion】Feeding of H. armigera larvae was deterred by piperine and sanshool via gustatory post-ingestive effect. No significant gustatory habituation occurred when the experienced stimuli paired with the control, and it seems that the binary mixture of piperine and sanshool could not mitigate the habituation. The two amides have a broad prospect in field application.