Scientia Agricultura Sinica

Study on the Characteristics of the Low-Molecular-Weight Glutenin Subunits in the Formation of the Wheat Varieties

YANG Rui, GAO Xiang, CHEN Qi-Jiao, LI Xiao-Yan, DONG Jian, MENG Min, ZHAO Wan-Chun, SHI Yin-Gang, CHEN Liang-Guo

Scientia Agricultura Sinica. 2013, 46(9): 1745-1755. doi:10.3864/j.issn.0578-1752.2013.09.001

Abstract ( 659 )

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【Objective】To understand the diversity origin of low-molecular-weight glutenin subunits (LMW-GS), and provide references for wheat quality improvements, the variation features of LMW-GS in the formation of wheat cultivars were characterized.【Method】With the capillary electrophoresis (CE), the seeds of three sets of hybrid materials (F5/F6), together with their corresponding parents, were used to test the variability of the LMW-GS. A series of specific primers, designed according to the genetic sequence published in GenBank, PCR amplification, gene cloning, and sequence analysis were employed to characterize the features of LMW-GSs. 【Result】The results of CE indicated that all the tested lines contained the relatively stable numbers of LMW-GS compared to the parents, while the accumulation amount of several subunits increased significantly. On the basis of the sequence analysis, all of the 54 genes (GenBank access numbers from KC222070 to KC222121, and from KC478714 to KC478715) belonged to the lmw-gs gene family because of their typical structure of LMW-GS, including the relatively conserved signal peptides, intermediate repeating regions with abundant variations and the conserved C-terminal domain. The similarity alignment between the advanced lines and their parents revealed that all the three sets of hybrid materials shared the extremely conserved sequences with their relevant parental lines and all these common sequences belonged to LMW-m subunits. The relatively conserved sequences originated from SNPs were the major proportion and both parents have been acted as their donors. It was also found the fine procedure of the extra Cys residue in the LMW-GS during the formation of wheat cultivars. What’s more, LMW-GS with 10 or 11 Cys residues was also observed in the present study. The variation of Cys residues mainly occurred in the C-terminal region Ⅱ, while only one variation in the C-terminal region Ⅲ. 【Conclusion】 During the process of wheat breeding, the m-type LMW-GS from paternal sources tend to be more conservative than other kinds of subunits, and SNPs play an important effect on the diversity of LMW-GS.

Mechanism Analysis of AtBT4 from Arabidopsis thaliana Against Botrytis cinerea

HAO Cong-Cong, YANG Ping, CHEN Zhan, JIA Jiao, ZHAO Bin, SI He-Long, HAN Jian-Min, XING Ji-Hong, DONG Jin-Gao

Scientia Agricultura Sinica. 2013, 46(9): 1756-1761. doi:10.3864/j.issn.0578-1752.2013.09.002

Abstract ( 738 )

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【Objective】The objective of this study is to reveal the interactions between AtBT4 gene and SA, JA signal pathways in Arabidopsis thaliana resistance to Botrytis cinerea.【Method】RT-PCR technology was used to analyze the expression of AtBT4 in Col-0 treated with SA, its analogues BTH, JA, ACC and B. cinerea, the expression of AtBT4 in various signals pathway mutants following SA or JA treatment and the expression of resistance-related genes in Col-0, bt4 and AtBT4 complemented plants. 【Result】The expression of AtBT4 in Col-0 treated with JA and B. cinerea was significantly enhanced. But no significant difference was observed in AtBT4 expression between the JA-insensitive jar1 mutants treated for 0 and 12 h with JA. The AtBT4 expression was significantly down-regulated in the eds5, sid2, and npr1 mutants compared with the Col-0 plants. Compared with the Col-0 and AtBT4 complemented plants, the expression of PR1, PR4, PDF1.2 and BIK1 genes was significantly down-regulated in the bt4 mutants. 【Conclusion】 The AtBT4 gene expression is induced by JA and B. cinerea and the AtBT4 gene deficiency alters expression of resistance-related genes, PR1, PR4, PDF1.2 and BIK1, suggesting the AtBT4 gene may be involved in the regulation of SA and JA singnal pathway to impact the resistance to B. cinerea in A. thaliana.

Screening and Identification of Sorghum Cultivars for Alkali Tolerance During Germination

LI Feng-Xian, ZHOU Yu-Fei, WANG Yi-Tao, SUN Lu, BAI Wei, YAN Tong, XU Wen-Juan, HUANG Rui-Dong

Scientia Agricultura Sinica. 2013, 46(9): 1762-1771. doi:10.3864/j.issn.0578-1752.2013.09.003

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【Objective】The objective of the current study was to explore the response of 35 different sorghum cultivars to alkali stress, and screen them for suitability for growing in alkaline soil and provide some guidance for their management under alkali stress.【Method】Mixed alkali were used (NaHCO3﹕Na2CO3 with the mole ratio of 9﹕1) to simulate the typical alkali stress environment. The 35 sorghum cultivars were treated with 25 mmol•L-1 solution or distilled water and incubated in Petri dishes for 10 days at humidity of 60%, light/dark of 12 h/12 h, illumination of 34 μmol•m-2•s-1 and 28℃/25℃ of day/night during germination. The germination force was measured on the fourth day and the germination percentage, leaf length, root length, leaf fresh weight, root fresh weight, leaf dry weight, root dry weight and remaining seed dry weight were measured on the tenth day. Through the fuzzy mathematics method of membership function, principal component analysis (PCA) and clustering analysis (CA), the alkali resistance of different sorghum cultivars were evaluated.【Result】The PCA results showed that root length, germination index and leaf dry weight were the most significant factors and are recommended as the main indexes for identifing alkali tolerance of sorghum at germination. The cultivars were ranked based on the membership function, and three groups on alkali tolerance were sorted from the 35 sorghum cultivars according to the CA. The analysis showed that seven of the 35 cultivars were strongly tolerant of alkaline conditions, e. g. Siza25, that twenty-two were had medium tolerance, e. g. Jiza 319 and Chiza 28, and that six were sensitive, e. g. Longza 9.【Conclusion】Principal component analysis showed that root length, germination index and leaf dry weight were the most significant factors and these are recommended as the main indexes to identify alkali tolerance of sorghum at germination, and in plant breeding and evaluation projects should be supported by CA and membership function to provide rigorous assessment of cultivars tolerance of alkalinity.

Impact of Shading on Growth, Development and Physiological Characteristics of Trollius chinensis Bunge

吕Jin-Hui , LI Yan-Feng, WANG Xuan, REN Lei, FENG Yan-Meng, ZHAO Xia-Lu, ZHANG Chun-Lai

Scientia Agricultura Sinica. 2013, 46(9): 1772-1780. doi:10.3864/j.issn.0578-1752.2013.09.004

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【Objective】This study investigated the effects of shading treatment on growth, development and physiological response of Trollius chinensis Bunge in order to determine the optimum light intensity and best growth period for shading treatment suitable for T. chinensis growth. 【Method】Two-year-old T. chinensis plants were subjected to shading treatments of 0, 40%, 60% and 80% of light intensity of sunshine, respectively, from 5th May to defoliation. Morphological indicators were recorded for plant height, the number of leaves, leaf area, petiole length, crown width, diameter of corolla and the number of flowers per plant. Physiological parameters were measured including the malondialdehyde (MDA) content, relative membrane conductivity, superoxide radical content, superoxide dismutase (SOD) activity, peroxidase (POD) activity, soluble protein content and chlorophyll content. The ultrastructure of leaf chloroplast derived from various shading treatments was observed by transmission electron microscopy. 【Result】 The activities of growth and development of T. chinensis were promoted by shading treatments, in which, 40% and 60% shading treatments prolonged the flowering periods, the number of flowers per plant and the diameter of corolla were also increased. The number of grana, grana thickness and the number of grana lamellae were increased significantly by 60% and 80% shading treatments, but the number of starch grains in chloroplast was significantly reduced by shading treatments. Physiological characteristics of T. chinensis varied between the duration/growth periods when shading treatments were applied. The plants showed a weak light stress response after shading treatment for 40 days. Following the increase on shading durations the chlorophyll content and SOD activity were decreased while MDA content and relative membrane conductivity were increased. Shading treatment for 80 days boosted the growth of T. chinensis. The relative conductivity and the superoxide radical were significantly lower than the control while the chlorophyll content was significantly higher than the control. The soluble protein content and the SOD activity were enhanced under 60% and 80% shading treatments. Shading treatment for 120 days inhibited plant growth, increased the relative conductivity significantly and decreased the POD activity .【Conclusion】 Full light and 40% shading treatment were beneficial to T. chinensis from May to the middle of June, but 40% and 60% shading treatments were good choice in summer. It would be advisable that there were no shading in late summer and early autumn.

Multi-Index Irrigation Information Management System Based on WebGIS

CHEN Zhi-Fang, WANG Jing-Lei, LIU Zu-Gui, SONG Ni

Scientia Agricultura Sinica. 2013, 46(9): 1781-1789. doi:10.3864/j.issn.0578-1752.2013.09.005

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【Objective】The objective of this study is to improve the precision of irrigation decision and high efficient utilization of water resources.【Method】Based on precise irrigation, a system for multi-index irrigation management was developed by using the principle and method of system engineering and software engineering, which mixed with various technologies, such as the computer, database, networks and geographic information system, etc. 【Result】Taking the crop water requirement as the foundation, the decision models of single factor and multiple factors were constructed through considering the influence of the soil, crop, field monitoring, irrigation and drainage information, meteorological factors and so on. The information data such as irrigation area, real-time meteorological factors, crop water requirements, effective rainfall, the decision results, and soil moisture distribution were published by using the function of SuperMap IS.net. The operation personnel could get the total situation of the irrigation district and the decision results through the map Also, the system could analyze and integrate the discrete data of irrigation district. 【Conclusion】 The system was applied in the irrigation district of the Renmin Shengli Canal and Guangli. The results showed that application of the system improved the irrigation management level and decision accuracy, and also realized the intelligence and information of irrigation management and decision.

Molecular Characterization of T-DNA Integration of the Ustilaginoidea virens Mutant 5062

YU Mi-Na, HU Jian-Kun, HUANG Lei, YU Jun-Jie, YIN Xiao-Le, NIE Ya-Feng, CHEN Zhi-Yi, LIU Yong-Feng

Scientia Agricultura Sinica. 2013, 46(9): 1790-1798. doi:10.3864/j.issn.0578-1752.2013.09.006

Abstract ( 736 )

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【Objective】 The objective of the experiment is to establish a method for cloning genes of the Ustilaginoidea virens mutant, and to shed light on the pathogenic mechanism of the U. virens, depending on a new mutant 5062, which has increased virulence. 【Method】With the wild type strain 70-22 as a control, phenotypic analysis, such as growth rate, colonial morphologies and sporulation, were analyzed. TAIL-PCR combined with RACE were used to identify the T-DNA integration site and the genes of flanking right site of the T-DNA. The expression levels of the genes were analyzed by RT-PCR as well. 【Result】Colonial morphologies and pigment on PSA and TB3 medium were no different between 70-22 and 5062. But on MM and water-agar medium, 5062 grew slower and produced more conidiophores. The flanking sequences of T-DNA were non-adjacent in the wild type. The expression of the gene flanking right site of the T-DNA was upregulated.【Conclusion】In 5062, both the T-DNA insertion and inversion of chromosomal fragment caused a mutation of the increased virulence.

Transformation of Agrobacterium tumefaciens-Mediated Colletotrichum gloeosporioides and Identification of Transformants

WANG Hai-Yan, LI Bao-Hua, ZHANG Qing-Ming, LI Gui-Fang, DONG Xiang-Li, WANG Cai-Xia

Scientia Agricultura Sinica. 2013, 46(9): 1799-1807. doi:10.3864/j.issn.0578-1752.2013.09.007

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【Objective】The objective of this study is to optimize the Agrobacterium tumefaciens-mediated transformation (ATMT) technology system of Colletotrichum gloeosporioides, screen and identify the transformants and compare the difference of biological characteristics and pathogenicity between wild-type strain and transformants.【Method】Using the conidia of C. gloeosporioides strain LXS010101 as transformation recipients, A. tumefaciens strain EHA105 carrying plasmid pBIG3C harboring the hygromycin B phosphotransferase gene (hph) was transformed into C. gloeosporioides. The obtained transformants were screened and identified by hygromycin B resistance, PCR and Southern blot analysis. Partial transformants were selected randomly and analyzed on the colony morphology, mycelia growth rate, conidia development and pathogenicity.【Result】Successful transformation of C. gloeosporioides was performed and the highest efficiency reached on 439 transformants per 1×105 spores. The optimal transformation conditions were that 1×105 spores per milliliter of C. gloeosporioides spore suspension were co-cultured with Agrobacterium cells at 22℃ for 24 h, in the presence of co-culture medium containing acetosyringone (AS) at 200 μmol•mL-1. The transformants were stable when grown on PDA medium without hygromycin B for five times and were verified by PCR amplification with the hph primers and by Southern blot analysis with the hph probe. The results showed that all the detected transformants could be amplified the target bands and all the T-DNA were single-copy inserted into the genome of C. gloeosporioides. Compared with the wild-type strain LXS010101, most of the transformants did not change on the colony morphologies, however, in which 23.33% of the transformants decreased the mycelia growth rate significantly and two strains ATJ-3 and ATJ-15 could not produce conidia. In the other 28 transformants, 39.29% of the strains reduced the spore germination rate and 25.00% of the strains changed appressorium formation. In addition, 11 transformants with decreased their pathogenicity were characterized and ATJ-19 even completely lost its ability to infect apple.【Conclusion】The successful optimization of ATMT system of C. gloeosporioides and partial transformants were analyzed primarily indicated the usefulness of this approach for functional genetic analysis and revealing the pathogenesis mechanism in this important pathogenic fungus.

Molecular Cloning, Sequence Analysis and Expression Pattern Detection of α-Tubulin Gene from Helicoverpa armigera (Hübner)

YAN Shuo, ZHU Jia-Lin, ZHU Wei-Long, PAN Li-Long, ZHANG Qing-Wen, LIU Xiao-Xia

Scientia Agricultura Sinica. 2013, 46(9): 1808-1817. doi:10.3864/j.issn.0578-1752.2013.09.008

Abstract ( 669 )

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【Objective】The objective of this study is to clone and analyze a novel cDNA, named as HeTubA, encoding the α-tubulin from Helicoverpa armigera (Hübner), and detect the relative expression levels of HeTubA and HeTubB by real time PCR (QRT-PCR). 【Method】The total RNA was extracted from H. armigera 3rd instar larvae, and α-tubulin gene was cloned by reverse transcription polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE). The relative expression levels of HeTubA and HeTubB were detected in different adult organs and developmental stages by QRT-PCR.【Result】Sequencing and structural analysis showed that the ORF of HeTubA was 1 353 bp in size, encoding 450 amino acid residues (GenBank accession number JQ069957). HeTubA contained conserved residues of α-tubulin. The homologue analysis revealed that HeTubA shared high identity with Xestia c-nigrum, Papilio xuthus and Bombyx mori α-tubulin, which indicated that insect α-tubulin was conserved in evolutional process. QRT-PCR revealed that HeTubA and HeTubB mRNA expression was neither adult organ-specific nor developmental-stage-specific. Relative expression levels of HeTubA and HeTubB were the highest in compound eye, the lowest in abdomen. Expression of HeTubA peaked at last larval instar and pupal development, whereas that of HeTubB peaked at last larval instar and adult development.【Conclusion】HeTubA was cloned and mRNA expression levels of HeTubA and HeTubB were detected. 3D structural of HeTubA was constructed. The study is helpful for further researches on the function of HeTubA and HeTubB and development of a new-type pesticide.

Response of the Denitrifying Bacterial nirS Gene Community to Nitrogen Fertilizer in Paddy Field

SONG Ya-Na, WU Ming-Ji, LIN Yan

Scientia Agricultura Sinica. 2013, 46(9): 1818-1826. doi:10.3864/j.issn.0578-1752.2013.09.009

Abstract ( 533 )

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【Objective】The aim of this study was to investigate the effect of the nitrogen input amount on diversity of denitrifying bacterial community in paddy soil by nirS gene based on field location test of rice. 【Method】 Composition and abundance of denitrifying bacterial community in paddy soil were studied with the aid of denaturing gradient gel electrophoresis, cloning sequencing and real-time PCR by assay nirS gene in the forth year of a field experiment.【Result】Shannon-Wiener index of denitrifying bacterial nirS gene community in paddy field was calculated according to the image of DGGE. In the three treatments of nitrogen fertilizer (N1: 75 kg N•hm-2, N2: 150 kg N•hm-2 and N3: 225 kg N•hm-2), the Shannon-Wiener index was higher than that in the treatment of CK (without fertilizer), and the differences reached a significant level at heading and maturate stages of rice (P＜0.05). Whereas, among treatments of N1, N2 and N3 only Shannon-Wiener index of N3 was markedly higher than that of N1 in surface soil (P＜0.05) at tillering and heading stages of rice. Redundancy discriminate analysis (RDA) showed that the denitrifying bacterial nirS gene community structure in surface soil or in root zone soil all markedly correlated to growth stages of rice (P=0.002, 0.002), while there was a significantly correalation (P=0.002) between different levels of nitrogen fertilizer treatments and denitrifying bacterial nirS gene community structure only in surface soil. Real-time PCR analysis of nirS genes copies manifested that the increase in nitrogen input amount enhanced the abundance of denitrifying bacteria. At heading or maturate stages of rice, there was a upward trend of CK＜N1＜N2＜N3 in nirS genes copies in surface soil or in root zone soil, especially in surface soil at heading stage of rice (P＜0.05). At the same time, 8 DNA sequences of nirS gene cloning from DGGE band were got and logged in GenBank (Accession Number: JX997923, JX997924, JX997926-JX997931). In addition, the yield of rice in treatments of N1, N2 or N3 was 59%, 92%, or 107% more than that of CK. NO3--N in surface soil or in root zone soil increased with the increase of nitrogen input amount.【Conclusion】The findings demonstrated that the increase in nitrogen input amount promoted the enhancement in abundance and Shannon-Wiener index of denitrifying bacterial nirS gene community in paddy field, especially in surface soil. Changes in community structure of denitrifying bacterial nirS gene among different levels of nitrogen fertilizer treatments also took on in the surface soil. So there is an obvious response of denitrifying bacterial nirS gene community to nitrogen input amount in surface soil of the paddy field.

Distribution Characteristics of Organic Carbon Fractions in Soil Aggregates Under Tea Plantation of Different Ages

ZHENG Zi-Cheng, LIU Min-Ying, LI Ting-Xuan

Scientia Agricultura Sinica. 2013, 46(9): 1827-1836. doi:10.3864/j.issn.0578-1752.2013.09.010

Abstract ( 642 )

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【Objective】The purpose of this study was to explore and learn about the soil organic carbon stability and carbon sequestration capacity.【Method】 Based on the method of field investigation and laboratory analysis, the distribution of organic carbon fractions in soil aggregates was studied under tea plantation of different ages.【Result】The content of total organic carbon (TOC), labile organic carbon (LOC), particulate organic carbon (POC), water soluble organic carbon (WSOC) and potential mineralized carbon (PMC) in soil aggregates comparatively increased as soil aggregates particles diameter reduced, the maximum value concentrated was found at ＜0.25 mm or 0.5-0.25 mm diameter. But the content of soil aggregates microbial biomass carbon(MBC) comparatively decreased when soil aggregates particles diameter reduced, the maximum value was found at 5-2 mm diameter. The proportion of organic carbon fractions to total organic carbon in soil aggregates decreased as soil aggregates particles diameter reduced. The content of TOC in soil aggregates increased when the age of tea plantation addition, the content of LOC and POC in soil aggregates first decreased and then increased as the age of tea plantation addition, the maximum value was found at tea plantation of 22 age. The content of WSOC, PMC and MBC in soil aggregates first increased and then decreased as the age of tea plantation increased, the maximum value was found at tea plantation of 30 age. The proportion of organic carbon fractions to total organic carbon in soil aggregates decreased as the increase of tea plantation age. The contents of TOC and organic carbon fractions in soil aggregates in 0-20 cm layer were greater than in 20-40 cm layer and the proportion of organic carbon fractions to total organic carbon in soil aggregates in 20-40 cm layer was greater than in 0-20 cm layer. 【Conclusion】 The stability of organic carbon was strong for the smaller particle size aggregates, and there was the critical age of plant tea for the soil organic carbon sequestration abilities. The carbon sink effect in 0-20 cm soil layer was stronger than that in 20-40 cm soil layer.

Increasing Effect of N, P and K Fertilizer on Rapeseed in Different Regions of China

LI Hui, MA Chang-Bao, LU Jian-Wei, LI Xiao-Kun, REN Tao, CONG Ri-Huan

Scientia Agricultura Sinica. 2013, 46(9): 1837-1847. doi:10.3864/j.issn.0578-1752.2013.09.011

Abstract ( 721 )

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【Objective】 Field experiments were carried out to study the yield-increasing effect of nitrogen (N), phosphorus (P2O5) and potassium (K2O) fertilization in various rapeseed regions in China. General situation and regional characteristics were studied to provide a theoretical basis for a regional recommended fertilization. 【Method】 Increased yield, increase rate and agronomic efficiency (AE) were calculated based on the data from 2 106 field experiments in China from 2005 to 2010, which were used to analyze the yield-increasing effect of N, P and K fertilizer in different rapeseed regions.【Result】99% of the rapeseed field experiments in China showed yield increment under N fertilization. The average of increased yield and rate were 1 044 kg•hm-2 (in the range of 500-1 500 kg•hm-2) and 87.4% (in the range of 5-100%), respectively. Similarly with N fertilization, around 94% of the rapeseed field experiments presented yield increase when P fertilizer was applied. The average of increased yield and rate were 634 kg•hm-2 and 39.9%, which were mainly distributed in the range of 250-1 000 kg•hm-2 and 5-40%, respectively. Also there was 88% of the rapeseed field experiments showed yield increase when K fertilizer was applied. The average of increased yield and rate were 420 kg•hm-2 and 22.9%, mainly distributed in the range of 200-1 000 kg•hm-2 and 10-40%, respectively. Every kilogram of N, P2O5 and K2O would increase 6.2, 7.8 and 5.4 kg rapeseed, respectively. The yield increasing effect of N and P fertilizer was better on winter rapeseed in lower part of the Yangtze river than other regions, the average of its increase rate was 128.8% and 51.3%, respectively. The yield-increasing effect of K fertilizer on winter rapeseed in middle part of the Yangtze river was the best, with the averaged increase rate of 24.6%. The yield-increasing effect of N, P and K fertilizer on spring rapeseed was lower, with the average increase rates of 42.6%, 31.6% and 21.3%, respectively.【Conclusion】The fertilization effect of N, P and K fertilizer on rapeseed yield was remarkable in China. The yield-increasing effect was followed as N＞P＞K fertilizer. Nitrogen was the main nutrient factor affecting the yield of rapeseed. The yield-increasing effect of N, P and K fertilizer on spring rapeseed was lower than that on winter rapeseed. The effect of N, P fertilizer on winter rapeseed was best in lower part of the Yangtze river whereas the effect of K fertilizer was best in middle part of the Yangtze river. As there were a few field experiments with no yield increment or irrational fertilizer application, we suggest that the amounts of regional recommended fertilizer should be improved to achieve the target of high yield and fertilization efficiency.

Relationship Between Cropland Soil Arthropods Community and Soil Properties in Black Soil Area

LU Ping, XU Yan-Peng, TAN Fei, YANG Zhong-Qi, LIN Ying-Hua

Scientia Agricultura Sinica. 2013, 46(9): 1848-1856. doi:10.3864/j.issn.0578-1752.2013.09.012

Abstract ( 668 )

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【Objective】The characteristic of cropland soil arthropods community and soil properties under the disturbance of exogenous carbon and nitrogen was studied, to elucidate the effect of those disturbances on soil animal community.【Method】Using hand-sorting, modified Tullgren and Baermann’s methods, the cropland soil arthropods were collected from nine treatments, separately, i.e. Control (CK), exogenous carbon (C0.5, C1.0, C1.5, C2.0) and exogenous nitrogen (N0.5, N1.0, N1.5, N2.0) in the black soil area of Jilin during May to September of 2011, and the physic and chemistry character of the soil was also analyzed.【Result】A total of 70 485 individuals (111 unidentified） were collected, which belonged to eight classes, twenty orders, three suborders and 87 super-families. The individuals, groups and diversity of cropland soil arthropods were significantly affected by the sampling time and exogenous carbon and nitrogen. To compare to the control treatment(CK), soil organic carbon, soil total nitrogen, the C/N value of soil, individuals, group and diversity showed an increase in application exogenous carbon, while only soil total nitrogen, cropland individuals of soil meso-micro arthropods in application exogenous nitrogen. The number of individuals and groups were significantly correlated to soil moisture, soil temperature, soil organic carbon and soil respiration. Isotomidae and Dolichopodidae were both positively correlated to soil temperature and soil total nitrogen, while Mesostigmata, Oribatida and Onychiuridae positively correlated to soil organic carbon and soil respiration. The Redundancy analysis (RDA) showed that the dominant groups were significant effected by soil moisture, soil temperature and the C/N value of soil. The soil properties were much well explained by the dominant of cropland soil arthropod groups.【Conclusion】The composition and diversity of cropland soil arthropods were related to the sampling time and the types of exogenous, and affected by soil moisture, soil temperature and C/N value of soil. The cropland soil arthropod was sensitively indicator to inflect the changes in soil properties.

Cloning, Sequence and Expression Analysis of NADP-Malic Enzyme Genes in Apple

DONG Qing-Long, WANG Hai-Rong, AN Miao, YU Xian-Mei, WANG Chang-Jun

Scientia Agricultura Sinica. 2013, 46(9): 1857-1866. doi:10.3864/j.issn.0578-1752.2013.09.013

Abstract ( 820 )

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【Objective】This study is aimed to characterize the genes of MdNADP-MEs involved in malic acid metabolism in Malus×domestica B. by gene cloning, sequence and expression of MdNADP-MEs in different tissues.【Method】 Three full-length cDNA sequences of MdNADP-ME1,2,3 were isolated by RT-PCR. The obtained cDNA sequences and the deduced amino acid sequences were analyzed with bioinformatics methods. qRT-PCR was used to assess the expression of MdNADP-MEs in different tissues.【Result】The sequencing results showed that three cDNAs (designated as MdNADP-ME1, 2 and 3; GenBank Accession No. JX971883, JX971884 and JX971885) were 1 512 bp, 1 782 bp and 1 926 bp. They contained an open reading frame (ORF) of 1 512 bp, 1 782 bp and 1 926 bp, respectively, and their ORFs encoded an protein with 503, 593 and 641 amino acids, respectively. Amino acid sequence and structure analysis indicated that MdNADP-MEs contained five conservative amino acid areas (motif I-V) and two functional structure domains: malic and NAD_bind_1_malic_enz. The result of phylogenetic analysis showed that MdNADP-ME1 and 2 belonged to cytosolic dicot NADP-ME (group I), MdNADP-ME3 belonged to plastidic dicot NADP-ME. qRT-PCR result showed that MdNADP-MEs were constitutively expressed in all examined tissues, but showed different expression levels.【Conclusion】MdNADP-ME1,2,3 belong to NADP-ME family, possess highly conserved structures, and show different expression patterns in different tissues of apple.

Comparison of Light Interception Ability and Growth and Yield of Different Apple Tree Shapes on Dwarf Rootstock

DONG Ran-Ran, AN Gui-Yang, ZHAO Zheng-Yang, MEI Li-Xin, LI Min-Min

Scientia Agricultura Sinica. 2013, 46(9): 1867-1873. doi:10.3864/j.issn.0578-1752.2013.09.014

Abstract ( 717 )

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【Objective】 The difference in distribution of relative light intensity and yield and growth condition were studied with six different apple tree shapes and two rootstocks on apple trees in order to understand the suitable tree shapes for apple dwarfing rootstock cultivation in Northwest-loess plateau. 【Method】The structure index of trees, relative radiation, fruit quality and yield were measured by using TES-1332A.【Result】Spindle trees were more vigorous than Y-trellis and V-trellis trees. Tall spindle/M26 was better in distribution of relative light intensity, and the canopy volume ratio of relative light intensity more than 30% was 79.6%, total number of shoot of tall spindle was the most among the six spindle shapes, the yield of saplings was the highest. The relative light intensity within canopy of V-trellis was just lower than Y-trellis and the yield of V-trellis was just lower than tall spindle. The relative light intensity within canopy over 30% of Y-trellis/M26 was 91.8%, but the total number of shoot and output of Y-trellis was lower. The mean fruit mass of Y-trellis, V-trellis, slender spindle and improved spindle was better than that of tall spindle and freedom spindle. The fruit quality of Y-trellis and V-trellis with higher fruit weight, content of soluble sugar and pericarp color saturation was better than those of other tree shapes.【Conclusion】Tall spindle/M26 had the highest yield and V-trellis was the second, the fruit quality of Y-trellis and V-trellis was the best, and tall spindle was the second.V-trellis has a great potential in yield.

Formation of Canopy Structure, Yield and Fruit Quality of ‘Fuji’ Apple with SH6 Dwarf Interstock from Juvenility to Fruiting Early Stage

ZHANG Qiang, WEI Qin-Ping, LIU Song-Zhong, WANG Xiao-Wei, SHANG Zhi-Hua, LU Jin-Jin

Scientia Agricultura Sinica. 2013, 46(9): 1874-1880. doi:10.3864/j.issn.0578-1752.2013.09.015

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【Objective】 The goal of this study was to survey the developments of canopy structure, yield, and fruit quality in ‘Tall Spindle’ shaped ‘Fuji’ apple with SH6 dwarf interstock from juvenility to fruiting early stage and to provide research data for growers in making proper training and pruning, precocity and productivity in high density apple planting systems. 【Method】 The ‘Fuji’ apple with SH6 dwarf interstock planted in spring 2005 was used to investigate the dynamic changes of tree growth, number and proportion of different branch lengths, yields, and fruit qualities from 2006 to 2011. 【Result】The tree height and trunk circumference were increased rapidly before third year, and the tree height and rate of canopy coverage were 3.05 m and 33.64%, respectively, in fourth year. The total amount of branch was increased along with growth of tree age during the second year to seventh year, and they were 265.4 and 457.4 thousands per hectare in fourth year and sixth year. The amount of good spur and branch in length greater than 30 centimeters were increased and decreased rapidly, separately, in third year and fourth year, the proportion of good spur and branch in length of 30-60 centimeters were balanced more than 30% and less than 10% after foufh year. The yields were 10.8, 22.9 and 39.6 t per hectare, respectively, in fourth, fifth and sixth year. The fruit mass, percentage of fruit surface with red color, and soluble solid content were 299.79 g, 97.56% and 14.60%, respectively, in seventh year. 【Conclusion】The primary factors of low yield that were the total amount of branch and rate of canopy coverage were insufficiency before the fourth growth season. The key technical measures for precocity, high yield and good quality of ‘Fuji’ apple with dwarf rootstock in tall spindle are that the tree height, rate of canopy coverage, total amount of branch, proportion of good spur, and proportion of long branch no less than 30 centimeters are more than 3.0m, 60%, 500-800 thousands per hectare, 30%-35%, and 5%-10% by increasing planting densities and improving pruning at the end of the third year or fourth year.

Cloning and Expression Analysis of Eukaryotic Translation Initiation Factor 4E Gene and Screening of the Interactive Protein from Chrysanthemum×morifolium

LOU Wang-Huai, AN Juan, SONG Ai-Ping, CHEN Su-Mei, JIANG Jia-Fu, CHEN Fa-Di, FANG Wei-Min, GUAN Zhi-Yong

Scientia Agricultura Sinica. 2013, 46(9): 1881-1891. doi:10.3864/j.issn.0578-1752.2013.09.016

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【Objective】In order to study the expression patterns and function of translation initiation factor 4E in the Chrysanthemum, the eukaryotic translation initiation factor 4E gene was cloned from Chrysanthemum× morifolium ‘Jinba’ and its expression was analyzed, and the proteins interacting with the CmeIF4E protein were screened subsequently.【Method】CmeIF4E gene was cloned from Chrysanthemum by RT-PCR and RACE, the primers were designed according to the conserve motifs of eIF4E from several plant species, whose sequence of eIF4E has been reported. The expression patterns of CmeIF4E was analyzed by quantitative real-time PCR and subcellular localization, then screened the interacting proteins via yeast two-hybrid system.【Result】The eIF4E gene cloned from Chrysanthemum is 914 bp in length, ORF 654 bp, encoding 218 amino acids, named CmeIF4E. The sequences were submitted into GenBank database with accession No. JQ904591. The amino acid sequence analysis showed that the genetic relationships between Chrysanthemum and lettuce were the closest, and this result was consistent with plant taxonomy. Quantitative real-time PCR analysis showed that the CmeIF4E gene expressed in all tissues of Chrysanthemum ‘jinba’, the expression level of CmeIF4E in the young roots was the highest, next by the leaves, and was the lowest in stems. The subcellular localization assay showed that CmeIF4E was expressed in the cell nucleus, cytoplasm and cell membrane. The interacting proteins of CmeIF4E were related to the pathways of translation and post-translational modifications, photosynthesis, stress tolerance and defense.【Conclusion】CmeIF4E was constitutively expressed in Chrysanthemum tissues, while transient expression showed that CmeIF4E protein was located in the nucleus, cytoplasm and cell membrane. The candidate proteins screened by Y2H system showed that CmeIF4E might play an important role in translation initiation, photosynthetic system, plant resistance defense. All these results provided an important basis for the further study of the role of the CmeIF4E protein in Chrysanthemum.

Determination of Flavonoids in 10 Pummelo and Pummelo Hybrid Fruits by Ultra Performance Liquid Chromatography

FANG Bo, ZHAO Qi-Yang, XI Wan-Peng, ZHOU Zhi-Qin, JIAO Bi-Ning

Scientia Agricultura Sinica. 2013, 46(9): 1892-1902. doi:10.3864/j.issn.0578-1752.2013.09.017

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【Objective】This study aimed to determine 11 flavonoids in 10 pummelos and pummelo hybrids fruits and analyze flavonoids types and content in varieties of fruits by ultra performance liquid chromatograph (UPLC) method.【Method】The separation was carried on a ACQUITY UPLC BEH C18 (2.1 mm×100 mm，1.7 μm, column temperature 35℃). Methanol (A)/0.1% formic acid/water (B) mixture was used as the mobile phase at a flow rate of 0.3 mL•min-1 through gradient elution: (1) 0-5 min, 90%-80% A; (2) 5-10 min, 80%-30% A; (3) 10-15 min, 30%-20% A; (4) 15-16 min, 20%-90% A. 【Result】 The flavonoids types and content in citrus fruit depend on genetic genotypes. Naringin is a major flavonoid in pummel. Naringin and neohesperidin are major flavonoids in grapefruit. Naringin content is higher in Riored fruit (Peel: 27.5 mg•g-1 DW, Pulp: 2.73 mg•g-1 DW) and neohesperidin content is higher in Cocktail (Peel: 14.93 mg•g-1 DW, Pulp: 0.70 mg•g-1 DW). The accumulation of flavonoids in hybrid fruits were affected by their parents.【Conclusion】UPLC method was effective in qualitative and quantitative analysis of flavonoids in citrus fruits. The flavonoids were abundant in pummelo fruits and the difference of flavonoids composition in diverse citrus fruits was significant.

Optimization of Incubation of Sperm and Lentivirus and Transgenic Pigs Production

CAI Wei-Guang, XI Qian-Yun, XIAO Min, GUAN Li-Zeng, MENG Fan-Ming, SHU Gang, ZHU Xiao-Tong, JIANG Qing-Yan, WU Zhen-Fang, ZHANG Yong-Liang

Scientia Agricultura Sinica. 2013, 46(9): 1903-1914. doi:10.3864/j.issn.0578-1752.2013.09.018

Abstract ( 725 )

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【Objective】A new method was developed for producing transgenic pig using incubation of lentivirus and sperm. 【Method】Effects of sperm number, amount of lentivirus, incubation time and incubation temperature on sperm activity were assessed by the orthogonality design. PCR and southern blotting were used to analyze the transgene.【Result】The optimized incubation was incubation of 100 µL lentivirus (5×105cfu) with 1.0×107 sperms per mL for 30 min at 17℃. Using this procedure sperms were artificially inseminated into sows. PCR was used to detect the offspring. Fourteen of 49 (14/49) piglets were PCR-positive. Four of 8 PCR-positive pitlets showed foreign gene intergration revealed by using southern blotting.【Conclusion】Incubation indexes of lentiviruse and sperm optimized in this study are that 100 µL lentivirus (5×105cfu) with 1.0×107 sperms per mL for 30 min at 17℃, and transgenic pigs were produced by using this procedure. Foreign gene intergration were revealed.

Study on the Relationship Between Polymorphism of RERG Gene and Growth Traits of Three Goats Breeds in Guizhou

CHEN Zhi, LUO Wei-Xing, LIU Ruo-Yu, CAI Hui-Fen, CHENG Jie, SONG Tao-Wei, ZHANG Yi-Yu

Scientia Agricultura Sinica. 2013, 46(9): 1915-1922. doi:10.3864/j.issn.0578-1752.2013.09.019

Abstract ( 593 )

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【Objective】This study aimed at researching polymorphism of the second, third and fourth exons of ras-related and estrogen-regulated growth inhibitor (RERG) gene in three indigenous Guizhou goats (Qianbei Ma goat, Guizhou White goat and Guizhou Black goat) and its association with growth traits was analyzed. 【Method】PCR-SSCP and PCR-direct sequencing were applied to detect the polymorphism sites and the general linear model was used to analyze its association with growth traits.【Result】Four consistent SNPs sites were found in the fourth exon of RERG gene in such three breeds. They are 56 bp C→G, 826 bp A→G, 1 434 bp T→C and 1 798 bp A→T. The following fixed effects model was employed in analyzing the growth traits in goat. The body height was significantly higher in individuals with genotype AB than AA and BB and genotype EF than EE and FF (P＜0.01). A significant difference was found in body weight between genotype CD and CC and DD (P＜0.05). The 1 798 bp (A/T) site did not appear significant difference. 【Conclusion】The 56 bp (C/G), 826 bp (A/G) and 1 434 bp (T/C) sites will provide a study basis between RERG genes and growth traits as a well as certain theoretical foundation for better feeding and genetic resources of indigenous goats.

Study on Fetal Skin Hair Follicle Development and Morphology of China Super-Fine Merino (Gansu Type)

WU Yu-Yu, YUE Yao-Jing, GUO Ting-Ting, WANG Tian-Xiang, GUO Jian, LI Gui-Ying, HAN Ji-Long, YANG Min, LIU Jian-Bin, SUN Xiao-Ping, LI Fan-Wen, HE Yu-Qin, YANG Bo-Hui

Scientia Agricultura Sinica. 2013, 46(9): 1923-1931. doi:10.3864/j.issn.0578-1752.2013.09.020

Abstract ( 701 )

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【Objective】This study aims to research the organizational structure of China Super-Fine Merino hair follicles and hair follicle morphogenesis process, which will lay a histological basis for molecular regulation mechanism of Super-fine Merino hair follicle development.【Method】Tissue frozen section was conducted by embryonic skin of China Super-Fine Merino and the results were taken photography under a microscope.【Result】Results showed that the follicle structure of Super-fine Merino sheep consists of connective tissue sheath, outer root sheath, inner root sheath, hair shaft and hair bulb sections, as other mammals. Primary hair follicle occurred on 87th day of fetal period, vesicle structure of the secondary follicle could be found at the bottom of the primary hair follicle. At gestational 138 d, the primary basic hair follicle was mature. From gestational 87 d to 147 d, the density of primary follicle was gradually decreased and the density of secondary follicle was increased as the gestational age growing, meanwhile the density of primary follicle reached its maximum value (232.8±12.44) (per mm2) at gestational 117 d. The gestational Secondary Follicles to Primary Follicles ratio (S/P ratio) reached the maximum (9.96) at gestational age 126 d.【Conclusion】Primary follicles began to occur at 84 d of gestational age, and secondary hair follicle began to occur at 87 d of gestational age, gestational age 102 d in the original the secondary follicles neck bulge began regeneration to secondary-derived follicles, most of primary follicles and some of secondary hair follicle had matured. The differentiation of secondary hair follicle is a key factor which affected the density of hair follicle; it can effectively increase the density of hair follicle and improve the quality of the wool.

Effects of Prion Protein on the Regulation of Classical and Alternative Activation of BV2 Microglia in vitro

FU Yong-Yao, SHI Fu-Shan, WANG Ji-Hong, YANG Li-Feng, ZHOU Xiang-Mei, YIN Xiao-Min, ZHAO De-Ming

Scientia Agricultura Sinica. 2013, 46(9): 1932-1938. doi:10.3864/j.issn.0578-1752.2013.09.021

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【Objective】 The purpose of this study is to investigate the effects of PrPC on various forms of microglial activation. 【Method】 BV2 microglia were treated, respectively, with IFN-γ, IL-4, or IL-10, and the mRNA expression of PRNP was examined by RT-PCR. Then the effects of si-RNA-mediated disruption of PRNP on different parameters of microglial activation in IFN-γ, IL-4, or IL-10-stimulated microglia were analyzed by RT-PCR and western-blot. 【Result】PRNP mRNA expression was invariably downregulated in microglia upon exposure to IFN-γ, IL-4, or IL-10. PRNP silencing prior to cytokines treatment reduced the responsiveness of microglia to INF-γ treatment, significantly altered IL-4-induced microglial activation phenotype, and had no effect on IL-10-induced microglial activation.【Conclusion】Together, these results support a role of PrPC in the modulation of the shift of microglia from a quiescent state to an activated phenotype and in the regulation of the microglial response during classical and alternative activation.

Expression of Ghrelin and GHS-R1a mRNA in the Bovine Oocytes During in Vitro Maturation

MI Yan, ZHANG Ling-Li, LI Hai-Jun, DU Chen-Guang, CAO Gui-Fang

Scientia Agricultura Sinica. 2013, 46(9): 1939-1945. doi:10.3864/j.issn.0578-1752.2013.09.022

Abstract ( 588 )

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【Objective】 The aim of this study was to investigate the possibility of Ghrelin and GHS-R1a mRNA expression in bovine oocytes produced in vitro and the relationship between FSH and the expression of Ghrelin and GHS-R1a mRNA. 【Method】Relative real time RT-PCR technology was applied to detect the pattern of Ghrelin and GHS-R1a mRNA expression in oocytes, and the effect of different concentrations FSH and the combination of FSH and FSH binding inhibitor on the expression of Ghrelin and GHS-R1a mRNA were observed. 【Result】 Relative real time RT-PCR experiments confirmed that the Ghrelin and GHS-R1a mRNA levels in oocyte varied depending on the developmental stage, with the significant decrease of Ghrelin and GHS-R1a mRNA expression at 8 h in oocyte maturation process. And addition of FSH inhibited the expression of Ghrelin and GHS-R1a mRNA within oocytes, and this inhibition effect could be attenuated by FSH binding inhibitor. 【Conclusion】 The expression of Ghrelin and GHS-R1a mRNA could be declined with the rising of FSH concentration during in vitro maturation of bovine oocytes.

Expression Profiling of Cotton (Gossypium hirsutum L.) Trihelix Genes Responsive to Abiotic Stresses

LI Yue, SUN Jie, XIE Zong-Ming, LI Quan-Sheng, SI Ai-Jun, CHEN Shou-Yi

Scientia Agricultura Sinica. 2013, 46(9): 1946-1955. doi:10.3864/j.issn.0578-1752.2013.09.023

Abstract ( 521 )

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【Objective】The objective of this study is to dissect cotton trihelix genes expression alteration responsive to different abiotic stresses, thus providing a foundation for stress-related genes cloning and elucidating the molecular mechanism of cotton stress-resistance.【Method】Semi-quantitative RT-PCR and real-time PCR were employed to reveal the expression profiling of 24 GhGTs under different abiotic stresses (200 mmol.L-1 NaCl, drought, 4℃) and exogenous hormone (100 μmol.L-1ABA) treatments.【Result】The results showed that the expression of 12 GhGT genes were up- and down-regulated by salinity and drought stresses, and that of 7 GhGT genes and 9 GhGT genes were up- and down-regulated by cold and ABA treatments, respectively. Moreover, three genes including GhGT2, GhGT18 and GhGT23 were response to all above treatments. 【Conclusion】The results suggested that GhGTs may play important roles in upland cotton plant abiotic stress adaption.

Scanning Electron Microscopy Observation on the Antennal Sensilla of Tetrastichus planipennisi (Hymenoptera: Eulophidae)

GAO Yu, WANG Zhi-Ying, ZHAO Hong-Ying, LIU Xin

Scientia Agricultura Sinica. 2013, 46(9): 1956-1964. doi:10.3864/j.issn.0578-1752.2013.09.024

Abstract ( 636 )

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【Objective】 The objective of this study is to define the external morphological characteristics of antennae and sensilla of Tetrastichus planipennisi (Hymenoptera, Eulophidae), a larval endoparasitoid of the emerald ash borer (EAB), Agrilus planipennis (Coleoptera, Buprestidae) in Northeast Asia. 【Method】 Morphological characteristics of the antenna and the type, morphology, amounts and distributions of antenna sensilla of T. planipennisi were examined and photographed with scanning electron microscope. The difference in sensilla between both sexes was also compared. 【Result】 The antenna of T. planipennisi was geniculate and composed of scape, pedicle and flagellum. The female adult flagellum had five segments and the male had four segments. Six types of antennal sensilla were indentified in the antenna, including two types of placoclea sensillum (PSⅠ, PSⅡ), three types of trichodea sensillum (TSⅠ, TSⅡ, TSⅢ), one finger-like sensillum, one chaetiea sensillum, one basiconic mastoid sensillum and Böhms bristles. Morever, PSⅠ distributed in male adult and TSⅡ and TSⅢ only distributed in female. Total number of trichodea sensillum in female antennae was significantly more than those in male. 【Conclusion】 The antenna of male and female adult T. planipennisi was both geniculate. The length of the antenna was about equal, but the types and distributions of the sensilla differed very much. There were sexual differences in the types, distributions and desities between male and female adults.

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