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    01 February 2014, Volume 47 Issue 3
    CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS
    Haplotypes and Expression Analysis of Salt-Tolerant Gene GmNcl1 in Soybean
    HE Wei-1, LIU Lin-2, GUAN Rong-Xia-2, QIU Li-Juan-2
    Scientia Agricultura Sinica. 2014, 47(3):  411-421.  doi:10.3864/j.issn.0578-1752.2014.03.001
    Abstract ( 528 )   HTML ( 3 )   PDF (715KB) ( 1141 )   Save
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    【Objective】The objective of the study is to confirm sequence polymorphisms of GmNcl1 gene and its differential pattern in stress. 【Method】Amino acid sequence of GmNCl1 was compared by BLASTP to find homologous genes. Fifty soybean varieties were used for the sequencing of GmNcl1 promoter and gene, and indicating of salt tolerance, and haplotype cluster and phylogenetic tree were analyzed by MAGE, looking for the correlation between salt-tolerance and sequence of GmNcl1. The expression pattern of GmNcl1 by real time PCR analysis in roots of salt-tolerant cultivar Tiefeng 8 and salt-sensitive cultivar 85-140 under a series of treatments including the protein inhibitors, pH, ABA treatments and salt, alkali and drought stress. 【Result】 GmNcl1 was homologous with Na +(K+)/H+ exchanger CHX. Sixteen single nucleotide polymorphism loci and two InDel loci were found in GmNcl1 gene sequence, including one G/A (salt-sensitive/salt-tolerant) base change in exon 3, which caused the nonsynonymous mutation from alanine to threonine. Eighteen polymorphism loci were composed into fourteen haplotypes, of which there were three salt-tolerant haplotypes and eleven salt-sensitive haplotypes. The haplotypes, GAGATATTC (salt-tolerant)/TTT ---- CT (salt-sensitive) composed of six loci were high-effectively distinguished between the salt tolerance and sensitivity. In the roots of soybean seedling, the expression of GmNcl1 gene was increased under the Na+/H+ antiporter inhibitor, amiloride, and unchanged under Ca2+-ATPase inhibitor, sodium vanadate. The expression of GmNcl1 was increased under alkaline pH, and showed fluctuations under acidic pH. The expression of GmNcl1 was also induced by ABA, and up-regulated by salt, alkali and drought stress. The greater stress, the more up-regulated expression of GmNcl1. Expression of GmNcl1 showed a similar trend in Tiefeng 8 and 85-140 under salt stress, but there was the greater increase in 85-140 than Tiefeng 8.【Conclusion】GmNcl1 was highly similar with Na+(K+)/H+ exchanger, its sequence polymorphism was related to salt-tolerance and sensitivity. GmNcl1 was inducted by ABA and GmNcl1 might be involved in the balance of the organism internal environment, and be a key factor in the stress response to salt, alkali and drought stress in soybean.
    Mapping of Sesame Waterlogging Tolerance QTL and Identification of Excellent Waterlogging Tolerant Germplasm
    ZHANG Yan-Xin, WANG Lin-Hai, LI Dong-Hua, GAO Yuan, 吕Hai-Xia , ZHANG Xiu-Rong
    Scientia Agricultura Sinica. 2014, 47(3):  422-430.  doi:10.3864/j.issn.0578-1752.2014.03.002
    Abstract ( 887 )   HTML ( 11 )   PDF (511KB) ( 1104 )   Save
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    【Objective】Sesame is extremely sensitive to waterlogging and it is the main factor affecting the development of sesame production and the improvement of sesame yield in China, however, waterlogging tolerance related molecular biology research foundation is weak, so far, domestic and foreign studies on sesame waterlogging tolerance gene mapping has not yet been reported. This study focuses on mapping QTL related with waterlogging tolerance based on a sesame recombinant inbred lines (RIL) population, developing molecular marker related with waterlogging tolerance combining with analysis on sesame core collections, and identifying waterlogging tolerant germplasm. 【Method】RIL population was generated from 6 selfing generations after hybridization between Zhongzhi No.13 with high tolerance and Yiyangbai with extreme sensitivity to waterlogging. Based on molecular data of 113 polymorphism markers on RIL population, a genetic linkage map was constructed using MapMaker/EXP. 3.0 software. In 2009 and 2010, RIL population was tested by artificial waterlogging stress at flowering stage both at Wuhan and Ezhou, and phenotypic data of each line were obtained, including percentage of regular plants and percentage of live plants. Analysis of variance (ANOVA) on phenotypic data was carried out by Microsoft Excel 2010 software, then QTLs were mapped by QTLNetwork 2.0 software, using composite interval mapping (CIM) method. Subsequently, sesame core collections were scanned by markers linked with major QTL, their waterlogging tolerance phenotypic data were also collected and analyzed, the effective marker related with waterlogging tolerance was detected. Excellent germplasms with waterlogging tolerance were obtained by repeated identifying of waterlogging tolerance phenotype, and combined with the molecular marker assisted selection. 【Result】The length of genetic linkage map constructed in this study was 592.4 cM, a total of 70 marker loci were grouped into 15 linkage groups (LG), the average distance between adjacent markers was 8.46 cM. Total of 6 QTL related with waterlogging tolerance at flowering stage were detected, which located on LG 7, 9, 13, and 15, explaining 5.67%-17.19% of the phenotype variation, their additive effect was 2.7190-9.7302. One of them explaining the most phenotype variation was qWH10CHL09, its additive effect was 3.9394, and the alleles from the parent Zhongzhi No.13 toward increasing the trait value, SSR marker ZM428 was closely linked with this QTL (genetic distance was only 0.7 cM). This marker was verified among 186 sesame core collections, results showed that there were significant (P = 0.0163) differences between phenotypes of 2 groups of accessions with 2 different genotypes of marker ZM428, therefore, marker ZM428 can be used as effective marker for marker-assisted selection (MAS). Eight excellent germplasm with tolerance to waterlogging were also identified from the sesame core collection, their percentage of regular plants after waterlogging stress were higher than 70% and percentage of live plants after waterlogging stress were higher than 80%. 【Conclusion】In this study, 6 QTL related with sesame waterlogging tolerance were detected, one of them explaining the most phenotype variation of 17.19%, one effective marker was identified, and 8 excellent germplasm with waterlogging tolerance were identified.
    TILLAGE & CULTIVATION·PHYSIOLOGY & BIOCHEMISTRY·AGRICULTURE INFORMATION TECHNOLOGY
    Research on Maize Leaf Recognition of Characteristics from Transmission Image Based on Machine Vision
    TANG Jun-1, DENG Li-Miao-2, CHEN Hui-1, LUAN Tao-1, MA Wen-Jie-1
    Scientia Agricultura Sinica. 2014, 47(3):  431-440.  doi:10.3864/j.issn.0578-1752.2014.03.003
    Abstract ( 494 )   HTML ( 5 )   PDF (648KB) ( 596 )   Save
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    【Objective】The purpose of the study was to create database of characteristics from maize leaf transmission images, analyze the rules of characteristics variation with maize varieties and the recognition results of different types of characteristics in order to provide a basis for further research of identifying maize varieties from leaf transmission image of different growth periods based on machine vision. 【Method】 Twenty-one common varieties of maize were selected as the research materials. The maize leaves at jointing stage, small bell stage, large bell stage and tasselling stage were collected. A total of 420 high quality transmission images of maize leaves were taken in lamp box. The software for characteristic extraction and recognition of maize leaves was designed and developed based on Matlab R2009a, which included image preprocessing module, characteristic extraction module, neural network recognition module and threshold selection module. The transmission images of maize leaves at jointing stage, small bell stage, large bell stage and tasselling stage were pre-processed by the software. Then 48 characteristics of color group, shape group and texture group were extracted from transmission images of maize leaf, and a total of 20 160 characteristic data. In order to study the rules of characteinristics variation with maize varieties, the coefficient of variation of 48 characteristics of leaf transmission image among different maize varieties were analyzed. In order to search the important characteristics with strong ability of identifying maize varieties from transmission images of leaves, the Artificial Neural Network was built and the recognition rate of single characteristics from different time were analyzed. In order to study the recognition results, the recognition rates of the three groups of characteristics and the group combinations of characteristics from different time were further analyzed. 【Result】 The results in 4 stages indicated that there were significant differences in the coefficient of variation of 3 groups of characteristics among different maize varieties. The differences were stable with the growth of maize. The coefficient of variation of color group was the highest, then the texture group and the third was the shape group. The results at 4 stages also indicated that there were significant differences among the recognition rates of 48 single characteristics. The recognition rates were between 9.52% and 29.33%. According to the recognition rates, the important characteristics were in the following order: the standard deviation of R, the minor axis length, the standard deviation of H, the diameter, the average value of H, the standard deviation of V, the standard deviation of B, the invariant moment 6, the eccentricity, the average value of S, the external convex polygon area, the average value of B, the smoothness, the kurtosis of V, and the standard deviation of S. Those average recognition rates was over 18%. The average recognition rate of the color group was the highest which was 86.76%, then the texture group which was 78.05%, and the third was the shape group which was 68.67%. The stability of recognition rates of the texture group was the highest, then the color group and the third was the shape group. The average recognition rate of combinations of shape group and color group was the highest which was 92.29%, then the combinations of color group and texture group which was 90.29%, and the third was the combinations of shape group and texture group which was 87.43%. At jointing stage, there was no obvious difference among the recognition rates of combinations of 3 groups. At small bell stage, the recognition rate of the combinations of color group and texture group rose sharply, while the recognition rate of the combinations of shape group and color group fell slightly, and the recognition rate of the combinations of shape group and texture group showed a marked decline, which still remained above 82%. There was no significant difference between recognition rates of the shape group and texture group and the color group and texture group at jointing stage, large bell stage and tasselling stage. And the recognition rate of combinations of the shape group and color group was always the highest. 【Conclusion】 This study has provided rich primary data on characteristics from maize leaf transmission images, and has provided a quantitative basis and a new method for further research of identifying maize varieties in different growth periods, which will play a good reference role in crop varieties identification.
    On Boll-Setting Optimization Theory for Cotton Cultivation and Its New Development
    DONG He-Zhong-1, MAO Shu-Chun-2, ZHANG Wang-Feng-3, CHEN De-Hua-4
    Scientia Agricultura Sinica. 2014, 47(3):  441-451.  doi:10.3864/j.issn.0578-1752.2014.03.004
    Abstract ( 546 )   HTML ( 4 )   PDF (589KB) ( 906 )   Save
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    After 60 years of research and production practice, China not only has established an intensive farming based technology system of cotton cultivation with distinctive Chinese characteristics, but also has founded a relatively complete theoretical system for China cotton cultivation. Such a theoretical system of cotton cultivation has made important contributions to enable China to be the first major cotton producing country in the world. Boll-setting optimization theory is the core of the theoretical system for high-yielding and fine-quality cotton production in China. The establishment, main contents and prospects of the boll-setting optimization theory are reviewed in this paper. Fiber yield and quality are a function of boll setting, which is significantly affected by boll-setting period, spatial position of the bolls, and the physiological state of the cotton plant. Optimization of cotton fruiting can be realized through formation of more bolls at the best boll-setting period, and in the best spatial position in a cotton plant with the most healthy physiological state. Peak boll-setting period is a stage with high photosynthetic efficiency for a cotton plant. Cotton fruiting can be further improved by synchronizing the periods of high photosynthetic efficiency of leaf canopy, peak boll-setting and ample light and heat resources. To optimize cotton fruiting, it is necessary, on the one hand, to stabilize or increase both the biological yield and economic index and on the other hand, to simultaneously increase the number of bolls per unit area and boll size through coordinating cultivar, environment and cultivation measures based on the dry matter accumulation and distribution features of high-yielding cotton. Photosynthetic products are the material basis that determine cotton yield. Dry matter accumulation at different growth stages directly affects boll setting in cotton. Optimization of cotton fruiting requires the maintenance of a higher dry matter accumulation after full bloom throughout the boll opening stage and a relatively high harvest index. It also requires configuration of an ideal plant architecture and optimal crop population structure through proactive and anticipatory control of cotton ontogeny. Maturity performance is termed as senescence performance of a cotton plant during boll opening. It includes performances of normal maturity, late maturity and premature senescence performance, respectively. Disorders of sink and source, as well as root and canopy often decreased root-sourced cytokinin levels and increased abscisic acid content, which might lead to early or delayed senescence-associated gene expression, finally leading to the formation of abnormal maturity performance (late maturity and premature senescence). To optimize the boll setting, it is necessary to well coordinate the relationship between the sink and source, root and canopy to realize the normal maturation of cotton plants. The establishment of boll-setting optimization theory not only promotes the development of science and technology of cotton cultivation in China, but also provides a strong theoretical support for the healthy development of China's cotton industry. It is believed that boll-setting optimization theory will remain the core of China’s theoretical system for high yield and quality cotton cultivation in the future. However, with the adoption and development of simplification, mechanization, sustainable and green production technology, cotton fruiting optimization theory will also experience further changes and development to better support the development of China's cotton production.
    Morphological Characters and Physiological Mechanisms of Cold Resistance of Winter Rapeseed in Northern China
    YANG Ning-Ning-1, 3 , SUN Wan-Cang-2, 3 , LIU Zi-Gang-1, 2 , 3 , SHI Peng-Hui-1, 3 , FANG Yan-1, 2 , 3 , WU Jun-Yan-2, 3 , ZENG Xiu-Cun-1, 3 , KONG De-Jing-1, 3 , LU Mei-Hong-1, 3 , WANG Yue-1, 3
    Scientia Agricultura Sinica. 2014, 47(3):  452-461.  doi:10.3864/j.issn.0578-1752.2014.03.005
    Abstract ( 594 )   HTML ( 2 )   PDF (864KB) ( 1142 )   Save
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    【Objective】Low temperature is one of the important factors affecting the growth of the northern winter rapeseed, so it is very essential to study the mechanism of the cold resistance of winter rapeseed. The aim of the paper was to study the cold resistance of winter rapeseed by morphological characters, physiological and anatomical characteristics, then evaluate an relationship between the indexes and cold resistance, select reliable index of the cold resistance of winter rapeseed and establish an evaluation model for cold resistance, which will provide a basis for germplasm discovery and identification of cold resistance in Northern China.【Method】Seven different cold resistant winter rapeseeds were treated under low temperature at 8-leaf stage, the growth habits and leaf anatomical characteristics were observed and the physiologic index, biochemical index, root diameter and root dry weight were determined. The cold resistance of these winter rapeseeds was analyzed and a evaluation model was built by main component analysis, regression analysis, cluster analysis and path analysis. 【Result】A negative correlation was presented between LT50 and wintering rate, POD activity at 5℃ to 0℃, CAT activity at 0℃ to -5℃, free proline content at 10℃ to 5℃ (X1), soluble protein content at 5℃ to 0℃ (X2), growth habits (X5), root diameter and underground dry matter (X6). A positive correlation was presented between LT50 and POD activity at 0℃ to -5℃, ratio of palisade tissue against spongy tissue (X3), thickness of palisade tissue, thickness of leaf, stomatal area (X4). 【Conclusion】The cold resistance order of 7 cultivars by principal component analysis was the same as the wintering rate. Six indexes were chosen by regression analysis to build mathematical evaluation model, i.e., Y=-7.58-0.018X1- 0.542X2+3.903 X3-0.002 X4+0.195X5-0.066X6, (R2=1.000). The materials were divided into five clases by cluster analysis, and for the winter rapeseed classification of strong cold resistance, it showed creeping growth, small ratio of palisade tissue against spongy tissue, small stomatal area, large dry weight of root, large increase of POD activity and SP content when it is cold. The result is consistent with the sort. The path analysis result indicates that soluble protein content is the direct factor for the cold resistance of winter rapeseed.
    PLANT PROTECTION
    Identification and Analysis of Magnaporthe oryzae Avirulence Genes in Liaoning Province
    WANG Shi-Wei-1, 2 , ZHENG Wen-Jing-2, ZHAO Jia-Ming-2, WEI Song-Hong-1, WANG Yan-1, ZHAO Bao-Hai-1, LIU Zhi-Heng-1
    Scientia Agricultura Sinica. 2014, 47(3):  462-472.  doi:10.3864/j.issn.0578-1752.2014.03.006
    Abstract ( 517 )   HTML ( 1 )   PDF (714KB) ( 869 )   Save
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    【Objective】The objective of this study is to identify avirulence genes (Avr-genes), which existed in the prevalent fungus of Magnaporthe oryzae, understand the distribution of Avr-genes in the epidemic strains of different regions, and to provide references for rice cultivars distribution.【Method】According to the sequences of 6 Avr-genes which had been cloned and were associated with the rice blast pathogenicity, the primers were designed to amplify the target sequences of the epidemic rice blast fungus in Liaoning Province. The DNA, extracted from the 26 strains of rice blast fungi, was used as a template for PCR amplification. By analyzing the PCR products by AGE (agarose gel electrophoresis) and sequencing, the sequences of base and amino acid for each Avr-gene were examined and compared. Some candidate primers were designed to check out the Avr-gene which had no PCR product.【Result】There was no PCR product when amplified Avr1-CO39, Avr-pia and Avr-pii by the primers, and there was also no PCR product by using the candidate primers, which showed that Avr1-CO39, Avr-pia and Avr-pii didn’t exist in most of the M. oryzae isolated from the main rice growing regions in Liaoning Province. On the contrary, the PCR products of AvrPiz-t, Avr-pik and Avr-pita were detected by AGE, and different genome types showed the 3 genes below were carried by the fungus in different mutated types and rates. AvrPiz-t related to the resistant genes including Pi2, Pi9 and Piz-t was detected in 22 M. oryzae strains, and the sequences of 21 strains were same to which exited in AvrPiz-t. The results of this experiment illustrated that the rice varieties carrying Pi2, Pi9 or Piz-t could have broad-spectrum disease resistance to rice blast fungi. The sequence of No. 16 strain was not the same as others, because it lost a single base C in the position of 192 bp, which caused the frame-shift mutation. The mutation led the AA (amino acid) sequence to terminate in position of 72nd AA, and made the secretory protein lose the function of avirulence. Avr-pik associated to the resistant genes like Pik, Pik-p, Pik-m and Pik-s were detected by AGE. The results showed that all the strains had the target bands. Their sequence showed that there were 4 genotypes of Avr-pik allele in the strains (B, D, F and G), and genotype D which carried by 12 strains could be recognized by rice blast disease resistance genes of Pik and its alleles as Pik-m and Pik-p. The 9 strains which carried genotype B of Avr-pik had been reported before, but its function of virulence had not been tested. The genotype F which exited in two strains was firstly found to be the new allele of Avr-pik, and they appeared in Dandong and Panjin, respectively. The specific point mutation, the 143 (A/G) of the base sequence, was the difference between genotype F and D. Genotype G which was also firstly detected was carried by the remaining 3 strains which only appeared in the area of Xinbin County in Fushun. The difference between genotype G and D was the point mutation of 168 (G/A) in the base sequence, which caused the protein translation early termination, therefore, genotype G lost its avirulent function. According to the AGE results, the same bands of Avr-pita were detected in all the 26 strains, and the sequences were tested and analyzed. The results showed that the genes in 26 strains had 5 genotypes of alleles, and there were several point mutations between the 5 alleles and the Avr-pita sequence, and the changes of the sequences caused the missense mutation. The difference of AA among the 5 genotypes were the 3 point mutation of 83 (D/N), 192 (Y/C) and 207 (K/R), which had been found in a structural domain of Avr-pita. The genotypes of allele all had been reported before. 【Conclusion】 The M. oryzae who carried Avr-pik, Avr-pita and AvrPiz-t distributed widely in rice growing regions of Liaoning Province. Breeding and cultivating the rice varieties carrying the corresponding resistance genes could alleviate the damage of the rice blast.
    Effects of Copper Stress on the Growth and Reproduction of Ostrinia furnacalis Guenée
    WANG Yu-Hong, LI Bao-Tong, TANG Li-Mei
    Scientia Agricultura Sinica. 2014, 47(3):  473-481.  doi:10.3864/j.issn.0578-1752.2014.03.007
    Abstract ( 511 )   HTML ( 3 )   PDF (552KB) ( 664 )   Save
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    【Objective】The objective of this study is to assess the effects of copper (Cu2+) stress on the growth and reproduction of Ostrinia furnacalis Guenée in the parental generation and its filial generation, and to provide a basis for the forecast of population dynamics of O. furnacalis and its control. 【Method】O. furnacalis larvae in three generations were continuously fed with standard artificial diet containing different concentrations of Cu2+ (25, 50, 100, 200 and 500 mg•kg-1) in the laboratory and the effects of copper (Cu2+) stress on the growth and reproduction of O. furnacalis were investigated.【Result】The larval survival rate, pupation rate and eclosion rate of O. furnacalis were significantly negatively correlated to the concentration of Cu2+ stress. The minimum concentrations of Cu2+ at 100, 50 and 50 mg•kg-1 were significantly decreased the survival rate and eclosion rate in the parental generation, the first and second filial generation, respectively, and 200, 50 and 50 mg•kg-1 were significantly decreased the pupation rate in each generation, respectively, which the larval mortality rates of the treatments at the concentrations of Cu2+ 200 mg•kg-1 and higher than 200 mg•kg-1 in the first and second filial generation were higher than 20%, the pupation rates of the treatment at the concentration of Cu2+ 500 mg•kg-1 in each generation were lower than 80%, the eclosion rate of the treatment was lower than 50% at the concentration of Cu2+ 200 mg•kg-1 in the second filial generation. The larval and pupal body mass of O. furnacalis were significantly negatively correlated to the concentration of Cu2+ stress. The minimum concentration of Cu2+ at 50 mg•kg-1 was significantly decreased the larval body mass in each generation, and 500, 50 and 50 mg•kg-1 were significantly decreased the pupal body mass in the parental generation, the first and second filial generation, respectively, which the body mass of the third and fifth instar larvae of the treatments at concentrations of Cu2+ 50 mg•kg-1 and higher than 50 mg•kg-1 in each generation was significantly lower than that of non-Cu2+ controls. The pupal body mass of the treatment at the concentration of Cu2+ 500 mg•kg-1 in the parental generation was significantly lower than that of non-Cu2+ controls, and the pupal body mass of the treatments at the other concentrations of Cu2+ stress was not significantly different compared with non-Cu2+ controls. The pupal body mass of the treatments at concentrations of Cu2+ 50 mg•kg-1 and higher than 50 mg•kg-1 in the first and second filial generation was significantly lower than that of non-Cu2+ controls, while the pupal body mass of the treatment at concentration of Cu2+ 25 mg•kg-1 was not significantly different compared with non-Cu2+ controls. The durations of larval and pupal stage of O. furnacalis in each generation were significantly positively correlated to the concentration of Cu2+ stress, which the durations of larval stage of the treatments at the concentrations of Cu2+ 50 mg•kg-1 and higher than 50 mg•kg-1 were higher than that of non-Cu2+ controls. The durations of pupal stage of treatments at concentrations of Cu2+ 200 mg•kg-1 and higher than 200 mg•kg-1 in the parental generation, at the concentrations of Cu2+ 100 mg•kg-1 and higher than 100 mg•kg-1 in the first filial generation, and at concentrations of Cu2+ 50 mg•kg-1 and higher than 50 mg•kg-1 in the second filial generation were significantly higher than that of non-Cu2+ controls. The oviposition number per female daily and fecundity of adult female in each generation were significantly negatively correlated to the concentration of Cu2+ stress, which the oviposition number per female daily of the treatments at the concentrations of Cu2+ 50 mg•kg-1 and higher than 50 mg•kg-1 in each generation was significantly lower that of non-Cu2+ controls, and the fecundity of adult female of all treatments under Cu2+ stress in each generation was significantly lower then that of non-Cu2+ controls. The effects of Cu2+ stress on average mass per one thousand eggs in each generation was not significantly different compared with non-Cu2+ controls, while its hatching rate of eggs was significantly negatively correlated to the concentration of Cu2+ stress, which hatching rate of eggs of the treatment at concentration of Cu2+ 100 mg•kg-1 in the first filial generation was only 66.88%. 【Conclusion】Continuous Cu2+ stress had significant inhibition effects on the growth and reproduction of O. furnacalis in the parental generation and its filial generations, and its inhibition effects were positively correlated with the concentrations of Cu2+ stress.
    Progress in Research of Cold Storage of Insect Parasitoids
    ZHAO Jing, WANG Su, GUO Xiao-Jun, GAO Xi-Wu, ZHANG Fan
    Scientia Agricultura Sinica. 2014, 47(3):  482-494.  doi:10.3864/j.issn.0578-1752.2014.03.008
    Abstract ( 737 )   HTML ( 12 )   PDF (553KB) ( 1062 )   Save
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    Article has been withdrawn by author. 2017.10.1

    SOIL & FERTILIZER·WATER-SAVING IRRIGATION·AGROECOLOGY & ENVIRONMENT
    A Research on Response of Enzyme Activities to Long-term Fertilization in Lime Concretion Black Soil
    CHEN Huan-1, LI Wei-1, ZHANG Cun-Ling-2, QIAO Yu-Qiang-1, DU Shi-Zhou-1, ZHAO Zhu-1, CAO Cheng-Fu-1
    Scientia Agricultura Sinica. 2014, 47(3):  495-502.  doi:10.3864/j.issn.0578-1752.2014.03.009
    Abstract ( 517 )   HTML ( 6 )   PDF (590KB) ( 722 )   Save
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    【Objective】In order to provide a theoretical support for improving soil biochemical environment and soil quality, insuring high and steady yield of crops in view of the characters of “drought, waterlogging, concretion, leanness” in lime concretion black soil, the responsive mechanism of soil enzyme activities to fertilization in black soil was studied by the way of exploring effects of long-term fertilization on soil enzyme activities. 【Method】 Based on a 31 yeas long-term fertilization experiment in Yangliu of Anhui province, the change trend of activities of urease, acid phosphatase, neutral phospahtase, catalase and invertase in tilth soil (0-20 cm) was investigated with 5 patterns of fertilization (non-fertilization, application of single chemical fertilizer, application of single organic fertilizer, mixed application of organic and chemical fertilizer (with the same amount of nitrogen), mixed application of organic and chemical fertilizer (with the lager amount of nitrogen) in wheat-maize rotation system, as well as the correlation and paired comparison was analyzed to interpret the relationship among soil enzymes and seasonal variation, respectively. 【Result】 Compared with control, the activities of urease, catalase, and invertase increased significantly by 130.5% and 129.5%, 11.4% and 14.9%, and 31.4% and 12.0%, respectively, after wheat and maize harvested in the situation of long-term application of single organic fertilizer, while acid and neutral phosphatase were less affected. Long-term application of single chemical fertilizer effectively enhanced soil acid and neutral phosphatase activities, whereas other 3 enzymes activities were significantly inhibited in the same time. With the same amount of nitrogen, the activities of all the 5 enzymes were maintained at a higher level under mixed application of organic and chemical fertilizer, in which low activity of certain enzyme caused by unbalance fertilizing could be avoided. With the lager amount of nitrogen, mixed application of organic and chemical fertilizer increased acid and neutral phosphatase activities significantly, but the effect on urease, catalase and invertase activities was insignificant. There were mutual stimulations between urease and catalase, urease and invertase, catalase and invertase, though phosphatase had a negative correlation with other enzymes. In addition, the seasonal variation of urease, acid and neutral phosphatase, catalase, invertase activities in lime concretion black soil was significantly different in 5 fertilizer applications treatments. 【Conclusion】 With the same amount of nitrogen, mixed application of organic and chemical fertilizer was superior to application of single chemical or single organic fertilizer which made soil enzymes activities stay at a higher level and then will be beneficial to improvement of soil biochemical environment. The correlation of urease and catalase, urease and invertase, catalase and invertase was positive, but the acid and neutral phosphatase was negatively correlated with other enzymes. Seasonal variation of enzyme activities varied with types of enzyme and fertilizations.
    Effect of Crystal Structures of Potassium-Bearing Minerals on Aspergillus niger Growth Metabolism and Potassium and Silicon Release
    SUN De-Si-1, YIN Jian-Mei-1, CHEN Ye-1, 2 , CAO Fei-1, 2
    Scientia Agricultura Sinica. 2014, 47(3):  503-513.  doi:10.3864/j.issn.0578-1752.2014.03.010
    Abstract ( 493 )   HTML ( 1 )   PDF (856KB) ( 612 )   Save
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    【Objective】Effects of Aspergillus niger on weathering of potassium-bearing minerals and leaching rates of potassium and silicon are closely related to the ability of producing acids and extracellular metabolites (mainly including polysacchrides and proteins) by it. Now little is known about the relationships between potassium or silicon leaching rate and mineral crystal structure, and about the relationships between mineral crystal structure and growth metabolism of fungi. A strain of A. niger was used to leach potassium and silicon, respectively, from orthoclase and muscovite. 【Method】 A strain of A. niger was cultivated in solid and liquid Czapek’s media containing bauxite, orthoclase and muscovite powders and free-mineral powder, respectively, in order to survey the growing characteristics of A. niger in different mineral environments. Bioleaching experiments of potassium-bearing minerals were carried out in shake flasks to investigate the effects of mineral weathering and extraction of potassium and silicon by the strain. By measuring the contents of polysaccharides, proteins, organic acids, K2O, SiO2 , and pH values in culture media, and by observing the change of mineral surfaces before and after bioleaching with X-ray diffraction and scanning electron microscope, the difference of A. niger’s growth and metabolism and ability of mineral bioleaching was analyzed, when A. niger was cultured in different media containing orthoclase and muscovite respectively. 【Result】 Potassium-bearing minerals with different crystal structures promoted A. niger’s growth and metabolism to different extents. The strain of A. niger reproduced and grew faster in media containing potassium-bearing minerals than in media containing free mineral and a small amount of potassium-bearing bauxite. A. niger cultivated in potassium-bearing solid media in growth process exhibited a more obvious chemotaxis of minerals than in free mineral and bauxite solid media, and more obvious in the muscovite medium than in the orthoclase medium. Meanwhile, A. niger’s mycelium-mineral aggregation was formed much more obviously in liquid media containing potassium-bearing minerals, especially containing muscovite powder, than in media containing free mineral and a small amount of potassium bearing bauxite. A. niger produced 1.45, 1.97, and 2.45 g•L-1 of mixed organic acids, and 5.11, 9.96, and 12.25 g•L-1 of polysaccharides, and 6.25, 13.78, and 16.97 g•L-1 of proteins, respectively, after culturing for 10 days in potassium-free Czapek’s medium, orthoclase medium and muscovite medium. The contents of K2O and SiO2 released from muscovite were 79.10 and 57.78 mg•L-1 higher than from orthoclase by A. niger. The SEM images and XRD patterns of muscovite and orthoclase powders before and after bioleaching showed that the surface of muscovite was etched much more obviously and formed much more amount of amorphous substance or imperfect crystal particles than that of orthoclase. The characteristic peaks reflecting muscovite were obviously weakened and some of them were faded away, but the characteristic peaks reflecting orthoclase were slightly enhanced in XRD patterns of muscovite containing a small amount of orthoclase or orthoclase containing a small amount of muscovite. 【Conclusion】 The difference of crystal structures of potassium-bearing minerals is one of important factors resulting in the difference of mycelium-mineral aggregation, metabolites and acidic microenvironment formed in the interaction between fungi and mineral, when A. niger is cultured in media containing different potassium-bearing minerals. And then, maybe, the difference is one of the key factors interfering with A. niger’s mineral weathering ability and extraction of potassium or silicon. Potassium-bearing minerals can excite and promote A. niger’s growth and metabolism in media containing free available potassium. Muscovite can promote A. niger to produce much more amount of organic acids, polysaccharides and proteins in comparison with orthoclase. A. niger can selectively decompose different minerals according to the stability of the mineral crystal structure in the bioleaching system containing mixed minerals. A. niger can give preference to weather muscovite, when orthoclase with framework structure and muscovite with stratified structure coexist in a same leaching system.
    Analyses on the Emergies of Multiple Compound Cropping Systems from Double-Cropping Paddy Fields in Jiangxi Province
    SUN Wei-Min-1, HUANG Guo-Qin-1, CHENG Jian-Feng-2, LIU Bin-Bin-1
    Scientia Agricultura Sinica. 2014, 47(3):  514-527.  doi:10.3864/j.issn.0578-1752.2014.03.011
    Abstract ( 405 )   HTML ( 2 )   PDF (774KB) ( 651 )   Save
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    【Objective】 Grain problem is always a hot issue concerned by the world. Seeking efficient cropping patterns, improving land utilization efficiency and enhancing grain production are the most urgent researches at present. From the results of related researches fields at home and abroad, multiple cropping patterns are the important approaches of utilizing natural resources and increasing crop production, so agriculture all over the world is developing toward the direction of multiple cropping patterns. In 1987, H T Odum, the famous ecologist in American, proposed the theory of emery analysis to explore the function principles and simulation technology of the complicated terrestrial ecosystems, and extended it to the ecological, environmental, social and economic systems involved in human. Since 1980’s, applying energy inputs and outputs to measure the good or bad of different ripe cycles and cropping patterns were focused and applied widely, which were beneficial for regulating the relationship between ecological environments and economic developments and had important practical values of scientific estimation and reasonable utilizations of natural resources. In the experiment, the resource utilizations, input and output benefits of multiple compound cropping systems from double-cropping paddy fields were clarified, which provided a theoretical foundation for the farming system reforms of winter compound cropping patterns in double-cropping paddy fields of south China and sustainable development of farmland ecosystems, the scientific and technological supports of moving forward roundly regional modern agriculture development and agricultural modernization constructions, reference frames of increasing grain production, farmers income and rural prosperity in the nation and regions. 【Method】 Energies of crop economic yields, photosynthetic productivities, solar energy utilization efficiencies, inputs, outputs, operating efficiencies and environmental loads of seven multiple cropping systems in Yujiang country of Jiangxi province were analyzed comprehensively with the theories and methods of emery analyses on the basis of data from field location experiments and related statistic yearbooks. 【Result】 The results showed that the energies of crops economic yields in winter multiple cropping were 217.57×106-229.7×106 kJ•hm-2 and higher than 213.5×106 kJ•hm-2 in winter fallow (T1); and energies of crops economic yields in the T4, T6 and T7 were significantly higher than those in the other treatments, the increasing percent of energy in T6 was 8.5% and the highest. Energy increasing percent of T6 was the highest in 2008-2010, which indicated that T6 had a better superiority and stability. Photosynthetic productivities (11.99-14.03 g•m-2•d-1) and solar energy utilization efficiencies (1.46%-1.70%) in winter multiple cropping were markedly significantly higher than those (10.55 g•m-2•d-1 and 1.28%, respectively) in T1, the average increasing ranges were 14.4%-34.8% with the highest increasing percent (34.8%) of T3, and the changes of photosynthetic productivities and solar energy utilization efficiencies were exactly alike. Energies of crops economic yields in winter multiple cropping rotations were 220.9×106-229.7×106 kJ•hm-2 and higher than 217.5×106 kJ•hm-2 in winter multiple continuous cropping (T2), and the increasing percent of energy in T6 was 6.1% and was the highest, but no significant difference was found among all treatments. Winter multiple cropping rotations reduced crop photosynthetic productivity to a certain extent, photosynthetic productivities (11.99-13.10 g•m-2•d-1) in winter multiple crop rotations were lower than 13.67 g•m-2•d-1 in T2 except the higher in T3, the average decreasing ranges were 4.2%-12.4% with the highest reduction of T4. The solar energy utilization efficiencies in T3 (1.70%) and T6 (1.67%) were significantly higher than those in the other treatments (1.46%-1.58%) which decreased with the highest reduction of T4 and T5 in comparison with T1. Emergy analyses indicated that the emergy inputs and outputs of all treatments existed obvious differences; the least of net loss of topsoil was 2.98×1016 sej in T1, the most was 3.83×1016 sej in T5; the least of industrial supplement emergy was 1.62×1017 sej in T1, the most was 2.98×1017 sej in T4 and nearly two times in T1; the least of organic emergy was 6.55×1015 sej in T1, the most was 1.19×1016 sej in T7 and nearly two times in T1; the least of output emergy was 1.39×1016 sej in T1, the most was 5.42×1016 sej in T6 and nearly four times in T1. Emergy input ratios (3.12-4.57) in winter multiple cropping were higher than 2.84 in T1 except the lower of 2.81 in T3, the increasing percents in T4 and T5 were 61.1% and 50.4%, respectively; but only emergy input ratio (0.13) in T4 was higher than 0.08 in T1, those (0.06-0.07) in the other treatments were lower. Emergy ratios of environmental resources to total inputs in the most treatments were 0.17-0.26 with the maximum reduction of T4 and lower than that in winter fallow, which showed that winter compound cropping patterns were favorable to protecting farmland environmental resources. Emergy ratio of unrenewalbe to total environmental resources in all winter compound cropping patterns were 0.10-0.15 and higher than that in T1, which suggested that planting winter crops mostly depended on the unrenewalbe environmental resources and could increase soil loss. Emergy ratio of industrial supplement to total inputs in all treatments exceeded 0.8, but had no obvious differences, which indicated that the productions of all cropping patterns depended on the industrial inputs and counted against agricultural sustainable development. 【Conclusion】In conclusion, planting winter crops in double-cropping paddy fields were better for increasing photosynthetic productivities and solar energy utilization efficiencies of paddy fields. Photosynthetic productivities and solar energy utilization efficiencies of planting economic crops except mixed green manure in winter multiple cropping rotations were lower than those in winter multiple continuous cropping. Planting broad bean and pea in double-cropping paddy fields in winter were more superior to fallow-double cropping rice system; but planting rape in double cropping rice system in winter was one of the high input and output patterns for sustainable developments; thus, planting rape in double cropping rice system in winter was the optimum multiple cropping.
    HORTICULTURE
    QTL Mapping and Analysis of the Important Agronomic Traits of Beijingjietou×Xishuangbanna Cucumber Recombinant Inbred Lines
    MA Zheng, BAO Kai-Liang, LI Lei, QIAN Chun-Tao, CHEN Jin-Feng
    Scientia Agricultura Sinica. 2014, 47(3):  528-536.  doi:10.3864/j.issn.0578-1752.2014.03.012
    Abstract ( 450 )   HTML ( 2 )   PDF (703KB) ( 866 )   Save
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    【Objective】 A genetic map was constructed using Xishuangbanna cucumber (Cucumis sativus L.var. xishuangbannanesis Qi et Yuan) recombinant lines and QTLs associated with yield, chlorophyll content, fruit related traits, the lateral branch number and the first lateral branch node were identified. These results will be beneficial and helpful to cucumber plant selection and high yield breeding.【Method】Using JoinMap 4.0 software, a genetic linkage map based on 995 SSR primers data from 124 F9 recombinant inbred lines, derived from Beijingjietou cucumber and Xishuangbanna cucumber by using single seed descend method, was constructed. Using WinQTLcart2.5 software, QTLs associated with 12 cucumber traits related to chlorophyll content, fruit lengths, fruit diameters, length of fruit for seed harvest, diameter of fruit for seed harvest, stalk lengths of fruit for seed harvest, and lateral branch, the first lateral branch node were mapped.【Result】The linkage map consisted of 137 SSR (simple sequence repeat) markers, 7 linkage groups that spanned 591.2 cM with an average distance of 4.32 cM. A total of twenty-nine QTLs for the 12 cucumber traits were detected, and 6, 7, 9 and 7 QTLs for chlorophyll content characters, commercial fruit, mature fruit and lateral branch traits were detected, respectively. These QTLs were mapped on chromosomes 1, 2, 3, 4, 6 and 7, respectively. The QTLs explained 5.30%-19.24% of the phenotypic variation. The minimum contribution rate of Ldr4.2 is 5.30%, the maximum contribution rate of Lbn1.2 is 19.24%.【Conclusion】The results of QTL analysis on the 12 cucumber traits will provide a basis for gene fine mapping to unravel the genetic basis of yield related traits in cucumber. Moreover, the obtained genetic linkage map of this RIL population could be used for analysis of other important cucumber traits.
    Effects of Sustained Weak Light on Seedling Growth and Photosynthetic Characteristics of Potato Seedlings
    QIN Yu-Zhi-1, XING Zheng-1, ZOU Jian-Feng-2, HE Chang-Zheng-1, LI Yan-Lin-1, XIONG Xing-Yao-1, 3
    Scientia Agricultura Sinica. 2014, 47(3):  537-545.  doi:10.3864/j.issn.0578-1752.2014.03.013
    Abstract ( 509 )   HTML ( 2 )   PDF (1003KB) ( 1277 )   Save
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    1College of Horticulture and Landscape, Hunan Agricultural University, Changsha 410128; 2Center of Analytical Service, Hunan Agricultural University, Changsha 410128; 3Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences, Beijing 100081) Abstract: 【Objective】 The aim of this study was to investigate the effects of sustained weak light on growing conditions and photosynthetic characteristics of potato leaves at the seedling stage and to offer a theoretical basis for screening and utilization of new resistant potato resources to weak light.【Method】 The original cultivated potato Yan (S. tuberosum subsp. andigena var. yanacochense) and cultivated potato Favorita (S. tuberosum) were grown in a growth chamber at 50 μmol•m-2•s-1 and 350 μmol•m-2•s-1 light, respectively, using potato tuber (50 g) growing in medium. After 1 month treatment, the growth of the test materials was measured. Plant phenotype, the chlorophyll content, net photosynthetic rate, photosynthetic curve, the CO2 response curve were tested by LI-COR 6400 portable photosynthesis system. The stomatal morphological characters were measured using JEM-1200EX scanning electron micrography. The ultrastructure of chloroplast was observed by transmission electron microscope using the blade section after uranium acetate-citrate double staining.【Result】There was a difference between the treatment and the control in leaf size, morphology of plant. Favorita had weak vigour, slender seedling with smaller pale green foliage. Foliage and branch induction of Yan were very difficult and grew extremely slowly in or after treatment. Sustained light treatment significantly reduced the apparent quantum yield (AQE), light saturation point (LSP), maximum net photosynthetic rate (Pnmax), CO2 saturation concentration (CSP), and the chlorophyll content efficiency, and improved apparent carboxylation rate (EC), and CO2 compensation concentration (CCP) simultaneously. The light compensation point (LCP) of Yan was activated in the treatment. While the LCP and EC of Favorita were reverse. The EC, CSP of Yan had no difference between the treatment and the control. Growing under weak light, the stomatal density, chloroplast number declined. The numbers of grana and grana lamellae increased in Favorita. On the contrary, the number of grana lamellae decreased in Yan. 【Conclusion】 Foliage differentiation of potato was hypersensitivity to light. There was a significant genotypic difference. The development and growth of the sensitive genotypes were hindered when plants acclimated to low light (50 μmol•m-2•s-1). The damage could not be resumed by restoring strong light later. The sustained weak light treatment at seedling stage depressed photosynthetic rate (Pn) and dark respiration rate (Rday). Saturation point of plant fell off along with the light intensities reduced. Following the dark respiration rate (Rday) being decreased, Favorita potato could adapt to lower light intensity by reducing the light compensation point. On the contrary, Yan could not adapt to lower light intensity. Appeared organic matter accumulated difficulties because of the improved light compensation point (LCP), the narrowing effective intensity range and the increased dark respiration rate (Rday). Under sustained weak light stress, the mesophyll cell arrangement was changed , and the stomatal density, the stomatal apparatus and numbers of chloroplasts decreased significantly, the stomatal apparatus length-width ratio had a tendency to increase, and chlorophyll composition ratio changed. Favorita potato improved the capture ability of effective light sources by increasing the number of chloroplast grana, grana lamella and chlorophyll b. Yan potato had subdued capturing ability of effective light and CO2 affinity because the formation of chloroplast grana was affected and the number of chloroplast grana, grana lamella and stomatal density decreased.
    Heterologous Expression of Taxadiene Synthase Gene in Ganoderma lucidum
    XIN Yan-Hua-1, XIAO Zhao-Yan-1, YOU Lin-Feng-1, GUO Li-Qiong-1, 2 , LIN Jun-Fang-1, 2
    Scientia Agricultura Sinica. 2014, 47(3):  546-552.  doi:10.3864/j.issn.0578-1752.2014.03.014
    Abstract ( 437 )   HTML ( 1 )   PDF (666KB) ( 664 )   Save
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    【Objective】The objective of the study is to lay a theoretical foundation for the expression and production of taxol or its precursor products by using Ganoderma lucidum as a novel bioreactor.【Method】 For preparation of the G. lucidum protoplast, mycelium were collected and washed twice with 0.6 mol•L-1 mannitol, and enzymed with 1% lywallzyme at 30℃ for 3 h. Protoplasts were collected and suspended with 0.6 mol•L-1 mannitol, and the final concentration was adjusted to 1.0×108 cells/mL. For transformation of the exogenous gene (ts), the plasmid pgGIgpd-TS (1.0 μg) and pBgGI-hph (1.0 μg) were added in G. lucidum protoplast culture. Incubation on ice was performed for 30 min. Subsequently 0.5 mL of PEG was added and incubated at room temperature for 20 min. The transformed protoplasts were inoculated on the medium without hygromycin and cultivated 5-7 days at 25℃. After the colonies grew up, they were covered with the PDA semisolid medium with hygromycin. Genomic DNA and RNA were extracted from the mycelium of regenerated G. lucidum for transformant analyses. PCR and RT-PCR analyses were carried out for detection of the ts gene in putative transformants. For analysis of taxa-4(5),11(12)-diene, samples were analyzed on GC linked to a mass spectrometer using split-less injection. Temperature of the column was initially set and maintained at 100℃ for 1 min and was then increased to 300℃ at a rate of 8.0℃•min-1. The column was then maintained at this temperature for 2 min. 【Result】The putative transformants were obtained on the selective plates with hygromycin at 25℃. No colonies appeared on the nontransformed plates. This result indicated that the hph gene was transformed into the G. lucidum and obtained preliminary expression. The results showed that 4 out of 20 putative transformants were integrated into both ts and hph genes,with a co-transformation efficiency was 20%. RT-PCR results verified that all the 4 transformants had obtained the transcription of taxadiene synthase gene. Crude extracts from the G. lucidum were analyzed. It was found a GC peak at 16.762 min which was absent in the wild type control. The mass fragmentation pattern of this peak had the same diagnostic ions with taxa-4(5),11(12)-diene. The taxadiene was detected in the mycelia of transformed G. lucidum strain by GC-MS analysis.【Conclusion】This is the first report of the production of taxa-4(5),11(12)-diene by engineering a novel transformation system G. lucidum with taxadiene synthase gene. The authors proposed that G. lucidum is a promising host for the biotechnological production of precursors of paclitaxel, and the results for the study of molecular biology of G. lucidum is important.
    STORAGE·FRESH-KEEPING·PROCESSING
    Study on Postharvest Physiology, Main Diseases and Storage Technology of Sweet Potato
    ZHANG You-Lin, ZHANG Run-Guang, WANG Xin-Teng
    Scientia Agricultura Sinica. 2014, 47(3):  553-563.  doi:10.3864/j.issn.0578-1752.2014.03.015
    Abstract ( 805 )   HTML ( 5 )   PDF (905KB) ( 815 )   Save
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    【Objective】The planting area of sweet potato is 6.7 million square hectometer and its annual output is 100 million tons in China. However, the mildew deterioration rate of sweet potato production is about 15% due to the improper storage. The respiration type, storage temperature and main storage diseases of postharvest sweet potato were studied. To provide a theoretical basis and technical parameters for sweet potato storage, the effects of different fresh-keeping technologies on storage physiology and mainly diseases of sweet potato were investigated in the paper.【Method】Qinshu5 sweet potato as a main cultivated variety in Shaanxi was stored at (10±0.5), (11±0.5), (12±0.5) and (13±0.5)℃, respectively. The respiration intensity was evaluated by air flow method, the storage temperature was determined by comparing the decay index, the storage disease pathogen was identified by Koch's validation, ITS sequence analysis and morphological characteristics, and the dosage of thiabendazole (TBZ) fumigation was determined by comparing the spore breeding situation. The common sweet potato fungicide mildothane powder was used as the control. The polygalacturonase (PG) activity was measured by DNS colorimetry, amylase activity and superoxide dismutase (SOD) activity were determined by reagent kit, ascorbic acid content was determined by 2, 6-dichloro phenol indophenol titration, starch content was determined by acid hydrolysis method, reducing sugar content was determined by DNS colorimetry, and original pectin content was determined by carbazole colorimetry. The changes of SOD activity, amylase activity, PG activity, starch content, reducing sugar content, ascorbic acid content and original pectin content of Qinshu 5 sweet potato treated with TBZ fumigation, pre-storage low temperature (3, 4, 5℃), plastic bag packaging during storage were measured. Decay index and commercial percentage were calculated during the late period of storage. The statistical software Excel 2010 was used to analyze the test data and Duncan’s method was applied for the multiple comparison. 【Result】 The respiration type of sweet potato was climacteric variant and the optimum storage temperature was 11℃. Alternaria bataticola Ikata was the main pathogen that led to black spot disease during storage. TBZ could inhibit the spore reproduction obviously. The treatment that postharvest sweet potato callused at room temperature, being pre-cooled at low temperature, fumigated by TBZ, packaged with plastic bags, and then stored at 11℃ can significantly restrained the activities of amylase and PG, kept the activity of SOD in a highly active state, reduced the rate of starch convert into sugar, maintained the ascorbic acid content at a high level, delayed the physiological aging process of postharvest sweet potato, and kept the original quality of sweet potato well.【Conclusion】The postharvest sweet potato was placed to callus for 5 days under natural conditions, treated with low temperature at 4℃ for 2 days, then the storage temperature went up slowly with 1℃ every day, until it came to 11℃ the long-term storage started. During the storage temperature rising, the sweet potato was treated with 4.5% TBZ fumigant at the dosage of 6 g•m-3 for 3 hours and stored in plastic bags, with the rotten index being 0.185 and the commercial percentage being 93% after 210 days of storage. Under the suitable temperature, the sweet potato was preserved well by the callus before storage, low temperature treatment, TBZ fumigation combined with plastic package. Meanwhile it had a good safety and hygiene.
    ANIMAL SCIENCE·VETERINARY SCIENCERE·SOURCE INSECT
    Genome-Wide Association Analyses for Musle pH 72 h Value and Meat Color Traits in Sutai Pigs
    ZHOU Li-Sheng, YANG Jie, LIU Xian-Xian, ZHANG Zhi-Yan, YANG Bin, MA Jun-Wu
    Scientia Agricultura Sinica. 2014, 47(3):  564-573.  doi:10.3864/j.issn.0578-1752.2014.03.016
    Abstract ( 475 )   HTML ( 3 )   PDF (939KB) ( 1151 )   Save
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    【Objective】 The objective of this study is to identify candidate genes and molecular markers associated with meat quality traits of Sutai pigs. 【Method】 Genome-wide association analysis was conducted by using Illumina 60K SNP Bead-chip genotypes of 150 Sutai pigs. Phenotypic data of each animal included postmortem 72 h muscle pH(pH 72 h), pH drop from postmortem 45 min to 72 h (pHdrop_45 min_72 h), 72 h Minolta a, b, L (ColorM_a72 h, ColorM_b72 h, ColorM_L72 h) and subjective color score (ColorScore_72 h) of longissimus dorsi (LM) and semimembranosus (SM) muscle. Quality control was carried out using PLINK v1.07. SNP markers were removed if they had genotype-missing rates > 0.03 or minor allele frequencies (MAF) < 0.05 or Hardy-Weinherg P≤10-5. Samples were removed on low (<90%) call rate. After the quality control, all 150 samples passed the filter and a final set of 43 760 SNPs were selected for GWAS. The association analyses were conducted using GenABEL in the R software. SNPs were individually tested for association with all studied traits using a generalized linear mixed model and the genome-wide significance threshold was determined by the Bonferroni method. The influence of population stratification was assessed by examining the distribution of test statistics generated from the thousands of association tests and assessing their deviation from the null distribution in a quantile-quantile (Q-Q) plot. 【Result】 The relationship between LM_pH72 h and SM_pH72 h, between LM_pHdrop_45 min_72 h and SM_pHdrop_45 min_72 h, between LM_ColorM_b72 h and SM_ColorM_b72 h, between LM_ColorM_L72 h and LM_ColorScore_72 h and between SM_ColorM_L72 h and SM_ColorScore_72 h were significant (P < 0.05). There was no clear overall systematic bias in all studied traits and no very strong stratification existed. In total, 39 SNPs reached Bonferroni chromosome-wise significance and fell into 20 genomic regions of approximately 10 Mb or less. Among them, 17 SNPs significantly associated with pH values were detected on the chromosomes 3, 4, 10, 14 and X, except one unmapped SNP ASGA0082337. Other 22 SNPs for meat color located on the chromosomes 1, 3, 7, 10, 12, 14 and 15. The SNPs related to LM_ColorM_a72 h, LM_ColorM_L72 h, LM_ColorScore_72 h, SM_ColorM_L72 h and SM_ColorScore_72 h were not detected. Two SNPs M1GA0020074 and MARC0028756 on the chromosome 14 were significantly associated with pHdrop_45 min_72 h in both LM and SM. Linkage disequilibrium (LD) analysis by using Haploview version 4.2 software showed that M1GA0020074 and MARC0028756 were in a haplotype block spanning 433 kb.【Conclusion】The loci on SSC10, SSC14 and SSC15 appeared to have pleiotropic effects on several traits. BNIP3, PRKG1, ADRB3 and other genes near the significant SNPs are candidate genes for these traits and merit further validation.
    Expression and Localization and Effect of Cdc42 Protein During the Maturation of Bovine Oocyte
    LIU Yang-1, YIN Feng-Xia-Yin-Feng-Xia-2, BAI Chun-Ling-Bai-Chun-Ling-2, WEI Zhu-Ying-Wei-Zhu-Ying-2, ZAN Lin-Sen-Zan-Lin-Sen-1, LI Guang-Peng-Li-Guang-Peng-2
    Scientia Agricultura Sinica. 2014, 47(3):  574-585.  doi:10.3864/j.issn.0578-1752.2014.03.017
    Abstract ( 440 )   HTML ( 2 )   PDF (824KB) ( 414 )   Save
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    【Objective】 This study is designed to ascertain the expression and localization of Cdc42 protein during bovine oocyte maturation, and to clarify the effect of Cdc42 inhibition on maturation of bovine oocytes. 【Method】 Bovine oocytes in vitro matured for different times (0-24 h) were collected. Western blotting and immunocytochemical technique were adopted to detect the expression and distribution of Cdc42 protein from oocytes collected at different stages of in vitro maturation. A pair of primers with restriction enzyme sites of KpnⅠand EcoRⅠat the ends of forward primer and reverse primer respectively, was designed based on the mRNA sequence of bovine Cdc42 which was obtained from Genbank (accession No.: NM_001046332.1). The CDS of bovine Cdc42 gene was amplified by PCR, and then ligated with pMD19T vector to construct pMD19T-Cdc42WT, the cloning vector wild type of Cdc42. A pair of site-directed mutagenic primers was used in a PCR to obtain pMD19T-Cdc42T17N, the cloning vector of the dominant negative mutant of Cdc42. Both pMD19T-Cdc42WT and pMD19T-Cdc42T17N were digested by restriction endonuclease to obtain the fragment Cdc42WT (775 bp) or Cdc42T17N (775 bp). The obtained fragments were ligated to the multiple cloning site of pcDNA3.1(+) to construct the prokaryotic expression vectors. The cRNA of Cdc42WT and Cdc42T17Nwas synthesized by in vitro transcription, and then micro injected into the cytoplasm of bovine GV stage oocytes. The maturation rate of injected oocytes was detected. 【Result】 The expression level of Cdc42 protein remained constant over the whole bovine oocytes maturation process, but the distribution pattern varied as the meiosis proceeded. The distribution pattern that Cdc42 protein gathering at the cortex region was hardly found in oocytes of germinal vesicle (GV) stage, but occurred frequently and remarkably in metaphase Ⅰ (MⅠ) oocytes. The distribution pattern that Cdc42 protein gathering at the cortex region nearing meiosis apparatus rather than the other region of cortex was found in the oocytes at the stage of pro-metaphase Ⅰ (pMⅠ), and growingly frequent till to the stage of metaphase Ⅱ(MⅡ). The percentage of the pattern that Cdc42 overlaps to spindle was higher in the oocytes at the stage of anaphase Ⅰ (AⅠ) to telophaseⅠ (TⅠ), but went down when it comes to the MⅡ stage. Fragments of 785 bp were obtained via amplifying Cdc42 gene from the total cDNA of bovine oocytes. The cloning vector of wild type Cdc42 and the dominant negative mutant pMD19T-Cdc42WT and pMD19T-Cdc42T17N accorded with anticipation through the identification of restriction endonuclease digestion and sequence alignment. So bovine wild type Cdc42 gene and mutant Cdc42T17N were cloned successfully. The constructed prokaryotic expression vectors pcDNA3.1 (+) -Cdc42WT and pcDNA3.1 (+) -Cdc42T17N accorded with anticipation through the identification of restriction endonuclease digestion and sequence alignment, RNA fragments of 3079 bases and 987 bases were obtained by in vitro transcription in which the vector pcDNA3.1 (+) -Cdc42WT and pcDNA3.1 (+) -Cdc42T17N were used as the templates. The maturation rate of Cdc42T17N injected group was lower compared with Cdc42WT injected group and non-injected group (P < 0.05). 【Conclusion】 It was concluded that Cdc42 protein might be accumulated in the cytoplasm of oocytes during the follicular development, and was functional at the region of cortex and spindle during miosis of bovine oocyte. The normal activity of Cdc42 was necessary to bovine oocyte maturation and extrusion of first polar body.
    A-FABP and MSTN Gene Expression Patterns and Their Correlation in Different Tissues of Goose During Forced-feeding Period
    WANG Bao-Wei, LIU Teng, GE Wen-Hua, ZHANG Ming-Ai, LI Zhen, YUE Bin, CHENG Fan-Sheng
    Scientia Agricultura Sinica. 2014, 47(3):  586-593.  doi:10.3864/j.issn.0578-1752.2014.03.018
    Abstract ( 398 )   HTML ( 1 )   PDF (638KB) ( 798 )   Save
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    【Objective】Researches on the expression pattern of fat deposition-related genes in geese tissue are meaningful for variety selection. The expression pattern and correlationship between adipocyte fatty acid-binding protein (A-FABP) gene and myostatin (MSTN) gene in different tissues of goose during forced-feeding period were studied. 【Method】 Two hundred Qing Nong Gray geese at the age of 10 weeks were forced-fed and liver, lung, spleen, kidney, chest, leg muscle and abdominal fat during different forced-feeding periods were sampled and total RNA were extracted. The A-FABP and MSTN gene expression pattern in liver, spleen, lung, kidney, chest muscle, leg muscle and abdominal fat of geese were also evaluated using Real Time PCR. Diethyl ether extraction and gas chromatographic methods were employed to detect the content of fat and fatty acids in different tissues. Moreover, the above correlation was evaluated. 【Result】 A-FABP and MSTN genes were expressed in all tissues. A-FABP gene expression in the liver, abdominal fat, leg and chest muscles showed an up-down trend during forced-feeding period with a maximum level (15.90±0.04, 12.72±0.12, 8.88±0.06 and 6.33±0.13) at 24, 12, 18 and 24 d, respectively. MSTN gene expression pattern in the leg muscle and chest muscle showed a similar trend, while no significant difference were observed in the liver, abdominal fat, lungs and kidney (P<0.01). A-FABP gene expression was closely related to liver weight, liver fat, total polyunsaturated fatty acids and total saturated fatty acids in goose liver. The correlation index were 0.576, 0.510, 0.463 and 0.501, respectively, with significant differences (P<0.01), while MSTN gene expression exhibited no significant coefficients (P>0.05). 【Conclusion】 During forced-feeding for 18-24 d, liver weight and the content of fat, total unsaturated fatty acids and total saturated fatty acids in the liver raised significantly. A-FABP and MSTN gene in different goose tissues showed different expression patterns and negative correlations were observed during forced-feeding period. A-FABP gene in goose liver plays a leading role in regulating foie gras traits.
    Characterization of Cytochrome P450 Genes Involving in Ecdysteroidogenesis in Silkworm (Bombyx mori)
    CHENG Dao-Jun, LI Zhi-Qing, MENG Meng, PENG Jian, QIAN Wen-Liang, KANG Li-Xia, XIA Qing-You
    Scientia Agricultura Sinica. 2014, 47(3):  594-604.  doi:10.3864/j.issn.0578-1752.2014.03.019
    Abstract ( 503 )   HTML ( 3 )   PDF (740KB) ( 712 )   Save
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    【Objective】Insect metamorphosis is initiated by ecdysone, which is synthesized in prothoracic gland through cytochrome P450 enzymes-catalyzed enzymatic reactions. Silkworm (Bombyx mori) is a silk-producing insect. Interrupting or delaying the pupa-adult transition in silkworm will help to reform the flow of silkworm cocoon-processing and to further improve the quality of silk. The objective of this study is to identify and characterize P450 genes involving in ecdysteroidogenesis in silkworm and to provide candidate genes for genetic controlling of silkworm metamorphosis. 【Method】 Ecdysteroidogenesis-related P450 genes were identified in silkworm and other insects through sequence homology search. Phylogenetic tree of insect ecdysteroidogenesis- related P450 genes was constructed by ClustalW program. The mRNA expression profiles of ecdysteroidogenesis-related P450 genes from silkworm were checked by microarray and RT-PCR analysis. RNAi experiment was used to examine the effects of down- regulation of expression level of ecdysteroidogenesis-related P450 genes on silkworm metamorphosis. 【Result】Four P450 genes involving in ecdysteroidogenesis, including Cyp306a1, Cyp302a1, Cyp315a1, and Cyp314a1, were identified in the silkworm genome. Comparative analysis revealed that each edysteroidogenesis-related P450 gene existed as a single copy in the surveyed insects and all its orthologs in insects were grouped together well in the phylogenetic tree, indicating edysteroidogenesis-related P450 genes and edysteroidogenesis pathway were very conserved during insect evolution. Spatio-temporal expression profiling found that Cyp302a1, Cyp315a1, and Cyp314a1 showed a high expression in the ovary, testis, and head of silkworm larvae and were continually expressed during the late stages of silkworm metamorphosis. Cyp314a1 expression could be highly detected before wandering, pupation, or eclosion, showing a consistency with the stage-specific changes of ecdysone pulses. RNAi of Cyp314a1 resulted in not only larval death before silkworm pupation but also developmental abnormality of adult ovary. Two key genes involving in ecdysone signaling, HR3 and Ftz-f1, were also down-regulated after Cyp314a1 RNAi.【Conclusion】Silkworm P450 genes involving in ecdysteroidogenesis are evolutionarily conserved and down-regulating the expression level of the selected P450 gene interrupts normal pupation, suggesting that ecdysteroidogenesis-related P450 genes might be used in altering genetically the progress of silkworm metamorphosis.
    RESEARCH NOTES
    Comprehensive Evaluation of Relationship Between Rootstocks and Yield and Quality in Grafting Tomato
    GAO Fang-Sheng-1, 2 , WANG Lei-2, 3 , XU Kun-2
    Scientia Agricultura Sinica. 2014, 47(3):  605-612.  doi:10.3864/j.issn.0578-1752.2014.03.020
    Abstract ( 473 )   HTML ( 1 )   PDF (578KB) ( 716 )   Save
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    【Objective】Grafting has been used in protected tomato to limit successive obstacle, a lot of studies have been done on resistance against environmental stress, disease and yield increase by grafting, however, the effects of grafting on vegetable quality were yet not systematical and far from being completed. Therefore, in order to further make clear biological effect of tomato grafting, comprehensive evaluation of the effects of grafting on yield and quality in greenhouse tomato was studied to provide a theoretical basis for application of grafting.【Method】The change characteristics of the seedling quality, plant growth vigor, yield and fruit quality were measured by the same scion grafted on ten different rootstocks and the self-root grafting scion ‘Jinpeng No.1’ as control, and effects of grafting on seedling quality and fruit quality were evaluated on the basis of membership values of seedling quality and fruit quality calculated by the method of membership function.【Result】There was a significant difference in seedling growth of grafting plants among different rootstocks, whose coefficient of variation of leaf area and dry matter of root were 18.24% and 15.21% respectively. These grafting seedlings with rootstock of ‘MIKADO’ and ‘Kagemusha’ had higher seedlings quality with method of membership function, but the grafting survival rates on all rootstocks were higher than 90%. Plant growth vigor and yield of grafting tomato were significantly different among different rootstocks. Although the average rate of increasing yield was 34.87%, the coefficient of variation of yield was 17.16%. Except for yield on ‘Vespa’, other grafted plant yields were significantly higher than that of CK, which the increasing yield rate of grafting on ‘MIKADO’ and ‘Magnet’ were 66.82% and 57.56%, respectively. Fruit rigidity was significantly improved and fruit index was changed with grafting, and fresh mass of single fruit of grafting tomatoes were higher than that of CK except for those on ‘Vespa’, ‘Anka-T’ and ‘Support’. Grafting has a significant effect on fruit nutrient quality of tomato, higher effect on Vitamine C, lower effect on soluble solid, whose coefficients of variation were 69.16% and 8.87%, respectively. Fruit dry matter and the contents of soluble solid, soluble protein and titratable acid in all grafting treatments were improved, whereas the change value and direction of other indexes, such as fruit index, Vc, lycopene, soluble sugar and ratio of sugar to acid were dissimilar. The fruit quality was evaluated synthetically by calculating membership values of fruit nutrient quality indexes with membership function, which showed that the membership values of all grafting fruits were higher than the values of CK, and thus confirmed that fruit quality was improved by grafting.【Conclusion】The grafting seedling quality, plant growth vigor, fruit characteristics, yield and quality were significantly different with different rootstocks. Among the ten rootstocks determined, grafted plant yields were significantly higher than that of CK except for that of ‘Vespa’, higher increasing yield rate of ‘MIKADO’ and ‘Magnet’ rootstocks. Although the effects of grafting on fruit quality indexes showed differences in connect with different rootstocks, fruit quality of all grafting unions was improved synthetically, especially higher fruit quality was observed on rootstocks of ‘Anka-T’, ‘Vespa’ and ‘Rootstock 001’.