Scientia Agricultura Sinica

Molecular Cloning and Expression Analysis of Three Chain Extension Genes Related to Fatty Acid Synthesis in Upland Cotton (Gossypium hirsutum L.)

LIU Li-12, ZHAO Peng-1, WANG Dan-1, LIU Zheng-Jie-1, WANG Yu-Mei-3, HUA Jin-Ping-1

Scientia Agricultura Sinica. 2013, 46(17): 3523-3533. doi:10.3864/j.issn.0578-1752.2013.17.001

Abstract ( 600 )

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【Objective】Anabolism of fatty acid controls the synthesis of fat and affects responses of plants to abiotic stress. The temporal-spatial expression of related genes directly decides the content of oil in seeds, and affects stress responses of plants associated with fatty acids. The objectives of this study are to clone the important genes related to fatty acid synthesis and to analyze the expression of genes in the process of fatty acid accumulation and the responses of gene expression to stress with the aim to provide candidate genes for high-oil cotton breeding and stress-resistance cotton breeding.【Method】Full length cDNA of GhKAR, GhHAD and GhENR were cloned by means of homologous cloning. Bioinformatics analyses were performed on the obtained genes to describe the characteristics of encoded proteins. The expression model of target genes in tissues, the levels of their temporal-spatial expression and the expression characteristics under abiotic stress were analyzed through RT-PCR.【Result】The cDNA of GhKAR, GhHAD and GhENR were 1 119 bp, 906 bp and 1 318 bp in length, respectively, and encoded 283, 221 and 394 amino acids. GhKAR and GhENR belonged to NADB Rosemann supergene family and GhHAD belonged to hotdog supergene family. The genes expressed variously in different tissues, such as root, stem, leaf and young ovule at different development stages. The expression level of young ovules in high-oil cotton material was slightly higher than that in low-oil cultivars. The oil accumulated majorly from 20DPA to 30DPA in both high-oil and low-oil cotton cultivars, and differed much after 30DPA to maturation. That is, all the 3 genes in high-oil cultivar increased steadily in expression after 20 DPA, and reached the maximum at 30DPA, and expressed at a low value in low-oil cultivars meanwhile. The expression level of GhHAD and GhENR in high-oil cultivars was twice more than that in low-oil ones. The expression level of GhKAR, GhHAD and GhENR induced differently by low temperature. And GhKAR and GhHAD expressed in up-regulation when induced by ABA, and could not be induced by MeJA. The expression level of GhENR was up-regulated at 2h after induction by MeJA, and went down-regulated gradually thereafter, but could not be induced by ABA.【Conclusion】Full length cDNA of GhKAR, GhHAD and GhENR were cloned. Three genes played an important role in oil accumulation in cotton seeds. The characteristics of gene expression inferred these 3 genes involved in responding to physiological stress.

Construction of Genetic Linkage Map Using SSR Molecular Markers in Azuki Bean (Vigna angularis Ohwi and Ohashi)

LUO Wan-Xia, ZHANG Li, YANG Kai, LI Yi-Song, ZHAO Bo, LI Ming, WAN Ping

Scientia Agricultura Sinica. 2013, 46(17): 3534-3544. doi:10.3864/j.issn.0578-1752.2013.17.002

Abstract ( 557 )

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【Objective】 The published SSR primers of cowpea，common bean and EST-SSR (expressed sequence tags) primers of common bean were located and integrated in azuki bean linkage group to construct a linkage map of Chinese azuki bean with azuki bean SSR primers as anchor markers. More practical molecular markers will be supplied to gene mapping, cloning and molecular marker-assisted selection of azuki bean. 【Method】 A total of 1 473 SSR and EST-SSR primer pairs, including 906 SSR primer pairs of cowpea, 123 SSR and 248 EST-SSR of common bean, 196 SSR primer pairs of azuki bean, were used for PCR amplification to screen polymorphic markers between cultivated azuki bean parents of HB801×AG109 and GM892×AG110. Their F2 segregating populations were tested with the polymorphic markers. 【Result】 An integrated genetic linkage map of azuki bean containing 145 SSR and EST-SSR markers was constructed. This molecular genetic linkage map is composed of 59 azuki bean SSR anchoring markers, newly mapped 63 cowpea SSR markers, 9 common bean SSR markers and 14 common bean EST-SSR markers, as well as a stem colour marker-the purple stem trait which is located on linkage group 9. Genetic distance of purple stem trait from CEDG022 and cbess058 molecular markers is 0.9 cM and 0.1 cM, respectively. The total length of the linkage map was 823 cM and covered 11 linkage groups. The average distance between markers was 5.64 cM. The average distance of each linkage group spanned 74.82 cM. The average number of markers was 13.27 for each of 11 chromosomes. The length of each linkage group ranged from 7 to 26 markers was from 49.1 cM to 125.6 cM. 【Conclusion】Molecular markers of the relative species were freshly introduced in azuki bean and increased density of its genetic linkage map. It is useful for gene mapping and cloning, molecular marker-assisted selection in azuki bean.

Effects of Different Bowl Types and Densities on Grain Yield, Characteristics of Photosynthesis and Dry Mass Production of Bowl Transplanted Japonica Super Rice

ZHOU Xing-Tao-12, ZHANG Hong-Cheng-1, XU Ke-1, GUO Bao-Wei-1, CHEN Hou-Cun-3, CAO Li-Qiang-1, DAI Qi-Gen-1, HUO Zhong-Yang-1, WEI Hai-Yan-1, LI Ming-Yin-1, LI Gui-Yun-3, WU Zhong-Hua-3

Scientia Agricultura Sinica. 2013, 46(17): 3545-3561. doi:10.3864/j.issn.0578-1752.2013.17.003

Abstract ( 472 )

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【Objective】This experiment seeks to maximize the potential of rice bowl seedling planting specifications and reasonable density profile, explore new ways of labor saving and accomplish super high yielding cultivation of rice.【Method】The experiment was conducted by raising seedling of combinings with 3-bowl, 2-bowl and a single bowl seedling discs and using Wuyunjing 24 and Nanjing 44 as materials. Five different rice transplanting densities were designed, and the new bowl seedling planting production potential and the reasonable density were studied. 【Result】The treatments of 3-bowl together with 15×104 hm-2 holes, 2-bowl together with 18×104 hm-2 holes and a single bowl with 24×104 hm-2 holes accomplished the highest yield level, while the 2-bowl together with 18×104 hm-2 holes yielded most among the three treatments. For the different types of bowl seedlings, dry matter accumulation, its proportion of distribution, crop growth rate from heading to maturity stage and grain yield, a pattern at the initial showed a trend of single bowl＞2-bowl＞3-bowl under the lower basic seedling condition of 36×104 hm-2, 2-bowl＞3-bowl and single bowl under the medium basic seedling condition of 72×104 hm-2, 2-bowl and 3-bowl＞single bowl under the larger basic seedling condition of 90×104 hm-2 to 108×104 hm-2. In the three maximum yield conditions, dry matter accumulation, its proportion of distribution and crop growth rate from heading to maturity stage showed a trend of 2-bowl＞3-bowl＞single bowl. In the same basic seedling conditions, the effective tiller percentage showed a trend of 3-bowl＞2-bowl＞single bowl while the decreasing rate of leaf area showed a trend of single bowl＞2-bowl＞3-bowl. Under the three maximum yield conditions, the effective tiller percentage showed a trend of 2-bowl＞3-bowl＞single bowl while the decreasing rate of leaf area showed a trend of single bowl＞3-bowl＞2-bowl. Under the same basic seedling conditions, with the exception of the basic seedling in a minimum of 36×104 hm-2, dry matter per stem and sheath at heading, waxy, maturing stages and its maximum export and transformation showed a trend of 2-bowl and 3-bowl＞single bowl.【Conclusion】Under the conditions of 2-bowl and 3-bowl seedling discs, the holes for transplanting rice reduced by nearly 1/3 to 1/2, the effective tiller percentage becomes higher while the decreasing rate of leaf area becomes lower than the single bowl transplanting disc, the photosynthesis and dry mass production’s advantage becomes more obvious after heading stage in the appropriate or larger basic seedling conditions. 2-bowl and 3-bowl seedling transplanting discs have a higher yield potential, especially for the 2-bowl transplanting disc.

Meta-Analysis of the Relationship Between Maize Crop Yield and Plant Density from 1950s to 2000s in China

YANG Jin-Zhong, CHEN Ming-Li, ZHANG Hong-Sheng

Scientia Agricultura Sinica. 2013, 46(17): 3562-3570. doi:10.3864/j.issn.0578-1752.2013.17.004

Abstract ( 684 )

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【Objective】 The objective of the study is to reveal the general characteristics of yield-density relationships of maize plant density experiments in China, and to evaluate the suitability of meta-analysis as a tool to study maize yield response to plant density. 【Method】Based on requirements for meta-analysis, a collection of historical maize plant density experiments from 1950s to 2000s in China was built with some 1 500 pairs of plant densities and their crop yields. All eligible data sets were subject to statistical analysis such as gradients, boundary curves and 2-D kernel density. 【Result】Variation coefficient was 20.4% for optimum density, 33.65% for the highest yield and 30.8% for yield per plant under optimum density. Relative importance of yield components varied with combination zones of plant density and yield per plant. Upper boundary yields as a function of plant density followed the equation of Y＝-0.0134x3＋3.15x, within plant density range of [0.99, 15.0] plant/m2. Upper boundary yields as two stage functions of yield per plant were that increased along Y1＝113.1x, then declined along Y2＝-69.84x＋33.87 when yield per plant is larger than 0.185 kg. Plant density range within which there is the highest possibility for the yield of 15 mg?hm-2 fell in [7.0, 9.7] plant/m2. The 90th quantile for yield loss percentage was 6.18% due to 15% offset from the optimal plant density, and the 90th quantile due to 1 plant/m2 offset was 0.88 t?hm-2. 【Conclusion】Meta-analysis facilitates the distillation and abstraction of bulk data from maize yield-density experiments with multiple views, and it is applicable to other crop science areas.

Anti-TMV Activity and Isolation of Active Ingredients in Fruit from Chaenomeles sinensis

LIANG Hong-Yan-1, RONG Xu-1, CHEN Pi-Ting-1, MA Zhi-Qing-12, ZHANG Xing-12

Scientia Agricultura Sinica. 2013, 46(17): 3571-3579. doi:10.3864/j.issn.0578-1752.2013.17.005

Abstract ( 515 )

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【Objective】In order to find new botanical antiviral compounds, antiviral ingredients of Chaenomeles sinensis (Thouin) Koehne was isolated and evaluated against Tobacco mosaic virus (TMV). 【Method】Anti-TMV ingredients were isolated with bioassay-guided fractionation methods. Bioassay against TMV infection and replication of raw extracts and monomeric compound were determined using local lesion and potted plant methods on Nicotiana glutinosa and N. tabacum. 【Result】 Ethanol extract of C. sinensis displayed a significant inactivation effect on TMV in N. glutinosa with inhibition ratio of 97.76% and protection effect on TMV infection on N. tabacum var. K326 with efficiency of 49.98% at the concentration of 10 mg•mL-1. Ethyl acetate extract, n-butyl alcohol extract and water extract from ethanol extractum of C. sinensis all inactivated significantly TMV and the inhibition ratios were 89.02%, 82.84% and 84.74%. Dipentyl phthalate, β-amyrin, (8E)-hexadecenoic acid and (8E)-nonadecenoic acid were isolated and identified through comparison with data obtained via spectroscopic methods, and from values reported in the literature. Among them, β-Amyrin, (8E)-hexadecenoic acid and (8E)-nonadecenoic acid showed similar or better anti-MTV activities than 20% moroxydine hydrochloride•copper acetate WP, and the inhibition ratios were 27.51%, 33.33% and 41.08% at the concentration of 1.0 mg•mL-1. 【Conclusion】C. sinensis can be used as a biological antiviral agent in tobacco production and β-amyrin, (8E)-hexadecenoic acid and (8E)-nonadecenoic acid are its antiviral ingredients.

The Expression Level of Alkaline Phosphatase (ALP1) in Cry1Ac-Resistant and Susceptible Cotton Bollworm Helicoverpa armigera Hübner

ZHANG Tao-12, ZHANG Li-Li-2, WEI Ji-Zhen-2, XIAO Yu-Tao-2, LIANG Ge-Mei-2, ZHOU Rui-Yang-1

Scientia Agricultura Sinica. 2013, 46(17): 3580-3586. doi:10.3864/j.issn.0578-1752.2013.17.006

Abstract ( 473 )

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【Objective】 The objective of this study is to provide a theoretical basis for further clarifying Cry1Ac resistance mechanism and constituting reasonable resistance management strategy, the difference of ALP1 expression level between Cry1Ac-resistant and susceptible Helicoverpa armigera, the effect of Cry1Ac toxin on ALP1 expression level in Cry1Ac-resistant larvae and the relationship between ALP1 and Cry1Ac-resistant strains were analyzed. 【Method】 The expression levels of ALP1 in susceptible and Cry1Ac-resistant H. armigera larvae feeding on artificial diet containing Cry1Ac and no-Cry1Ac were compared using real-time quantitative PCR analysis. 【Result】 The ALP1 could express in whole development period of H. armigera, the highest expression level occurred in larvae, while the lowest was in pupae, and there was also relatively high expression level in moth. The highest expression level of ALP1 was in midgut of H. armigera larvae, followed by hindgut, foregut, malpighian tubules and epidermis. Compared with the susceptible H. armigera, the expression of ALP1 obviously increased in resistant larvae which had moderate resistance to Cry1Ac. The ALP1 expression of the resistant larvae significantly increased compared with the susceptible strain after they were fed on the artificial diet containing Cry1Ac, while the difference of ALP1 expression in 2nd and 3rd instar larvae was not significant between resistant strain and susceptible strain after they were fed on the normal artificial diet. The expression of ALP1 in the 4th instar larval midgut of all resistant strains were obviously higher than that in susceptible strain, and the expression level of ALP1 appeared downtrend along with the resistance ration increased. 【Conclusion】 The changes of ALP1 expression in midgut of resistant H. armigera may have a relationship with its resistance to Cry1Ac and metabolism of Cry1Ac.

Prokaryotic Expression, Purification and Functional Activity Assay in vitro of Soluble Trehalse from Apolygus lucorum

TAN Yong-An, XIAO Liu-Bin, SUN Yang, BAI Li-Xin

Scientia Agricultura Sinica. 2013, 46(17): 3587-3593. doi:10.3864/j.issn.0578-1752.2013.17.007

Abstract ( 546 )

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【Objective】 The aims of this study are to obtain recombinant protein which has a trehalase enzyme activity and identify the best pH and the optimum temperature on the basis of cloned the soluble trehalase from Apolygus lucorum previous. This research expected to reveal the molecular mechanism of trehalase synthesis and offer the basic research of the development of trehalase inhibitor pesticide. 【Method】 The vector containing ALTre-1 gene was double enzyme restricted by Nde I and Not I, then the cDNA identified by sequencing was constructed to pET28a vector and transformed into BL21 of E. coli. The target recombinant protein was over expressed and been purified by using Ni-NTA agarose, and then its activity assay and enzymatic characteristics were studied. 【Result】 The ALTre-1 gene could over express in E. coli and the target recombinant protein which purified through Ni-NTA agarose had higher trehalase activity ((184.83±13.39)nmol•μg-1•min-1) by using trehalose as substrate, the suitable reaction temperature was 55℃ and the best pH was 7.0. 【Conclusion】 Recombinant protein with trehalase activity was obtained, and the suitable pH and temperature of the highest enzyme activity were 7.0 and 55℃, respectively.

Effects of Mixtures of Different Organic Materials on Soil Nutrient Content and Soil Biochemical Characteristics

SONG Meng-Ya-12, LI Zhong-Pei-23, LIU Ming-2, LIU Man-Qiang-1

Scientia Agricultura Sinica. 2013, 46(17): 3594-3603. doi:10.3864/j.issn.0578-1752.2013.17.008

Abstract ( 653 )

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【Objective】 The objective of this study is to explore the effects of single type of organic matter and their combinations with various proportions of different organic materials on soil properties. The results would also provide a scientific basis and practical guidance to optimal utilization of agricultural organic wastes and achieve the aim of optimal configuration and agricultural sustainability regarding to soil quality. 【Method】 In this study, three organic materials were used, including rice straw, peanut straw and pig manure. Based on the mass ratio, the single type of organic matter only, the combination of both (rice straw and peanut straw mixing at the ratio of 7﹕3 and 3﹕7, respectively) and three ones (1﹕1). All single and mixture treatments were amended with 1% organic matters of the soil mass and cultivated for 90 days. Thereafter, soil nutrient and biochemical properties were measured. 【Result】 The results showed that different organic materials had a significant impact on soil nutrients and biochemical characteristics. Overall, mixtures of different organic materials were more effective than only single one for improving soil properties measured. The mixture treatments on soil nutrient availability showed additional effects, i.e. the nutrient release derived from mixtures were more than weighted average values of two and three materials. The additional effects of the mixtures on the activities of urease and invertase were not significant, for example, the increased effects of SZ, SHZ and 3S7H mixtures on invertase were 1.46%, 0.18% and 5.97%, respectively. Carbon mineralization rate of the mixture treatments had significant negative additional effects with the treatments of SH, SZ and 3S7H showed 9.91%, 23.54% and 22.95% lower than the weighted average values, respectively. And the corresponding decreased effects for the CO2-C production rate were 24.56%, 16.47% and 18.18%, respectively. BIOLOG analysis showed that AWCD value and carbon utilization potential of the treatments S and 7S3H were the highest among all treatments. The metabolic quotients (qCO2) in the S and 7S3H treatments were higher than those of other treatments. The materials combination of nutrient release and microbial biomass carbon and nitrogen had stronger positive addition. The materials combination of mineralization of organic carbon and qCO2 had stronger negative addition. 【Conclusion】 Results showed that mixing organic materials could improve soil nutrient contents enhance as well as soil biochemical activity. Compared to only one type of organic matter, materials mixture could provide more resources and favorable habitat for soil biological development, leading to acceleration of decomposition and nutrient release and finally improvement of soil fertility.

Research on Soil Organic Carbon Storage Distribution in the Grassland Ecosystem

SA Ru-La-1, LI Jin-Xiang-2, HOU Xiang-Yang-1

Scientia Agricultura Sinica. 2013, 46(17): 3604-3614. doi:10.3864/j.issn.0578-1752.2013.17.009

Abstract ( 631 )

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【Objective】The distribution characteristics of soil organic carbon (SOC) storage were studied for the purpose to determine the reasonable grazing management way and provide a basis for the regional carbon estimation in grassland ecosystem. 【Method】The SOC storage of 0-100 cm soil depth was measured by using field method of stratified sampling in the typical steppe.【Result】The SOC storage was between 9.72-14.84 kg•m-2 in the three study plots. No significant difference was found in SOC storage in three study plots of spatial distance. The significant difference of SOC storage was found between four different grazing managements treatments in study area, and the order was moderate grazing (MG)＞light grazing (LG)＞heavy grazing (HG)＞control area (CK). The SOC storage decreased with the increasing of soil depth, and a significant difference was detected between SOC among different soil layers. A positive significant relationship was found between SOC storage and soil layer (P＜0.01), and the relationship could be described by using logarithmic and linear equations.【Conclusion】The SOC storage was relatively stable in the typical steppe, and presented a vertical descending characteristics. The effect of different grazing managements on SOC storage in the same vegetation type was significantly higher than the effect of spatial distance. Moderate grazing is benefit for carbon fixation of grassland ecosystem.

Progress and Prospect in Research of Ecological Compensation for Agriculture

YAN Li-Dong-1, TIAN Miao-12, HE Dong-Cai-2, YUAN Hao-1, DENG Yuan-Jian-1

Scientia Agricultura Sinica. 2013, 46(17): 3615-3625. doi:10.3864/j.issn.0578-1752.2013.17.010

Abstract ( 591 )

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Ecological compensation for agriculture, as a key to guarantee the sustainable development of ecological agriculture and balance the farmer’s benefits, its theoretical study and practical exploration have become a hot spot in the field of ecological compensation in recent years. By searching and studying of literatures on it, the author gives comparative analysis on the ecological compensation for agriculture research status, research features, the existing problems, and the trend of development. The results show that, most researches mainly concentrate on the definition of agro-ecological compensation, anatomy of the compensation theory basis, construction of compensatory mechanism, policy support, host-guest definition, compensation criterion, and compensation mode. Research methods are mainly based on the experience of other countries, such as combination of qualitative research and quantitative calculation, combination of theoretical research and empirical analysis. The definition of ecological compensation for agriculture and host-guest legal status are the open questions. New constructive method, space optimal development, and performance evaluation deepening are analyzed in the study of the trend of ecological compensation for agriculture.

Effects of Calcium on Physiological Characteristics and Main Mineral Elements Absorption of Garlic

LI He-1, LIU Shi-Qi-1, LIU Zhong-Liang-2, FENG Lei-1, LIU Jing-Kai-1, CHEN Xiang-Wei-1, WANG Yue-1

Scientia Agricultura Sinica. 2013, 46(17): 3626-3634. doi:10.3864/j.issn.0578-1752.2013.17.011

Abstract ( 551 )

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【Objective】The effects of different calcium levels on physiological characteristics and N, P, K, Ca, Mg absorption of garlic were studied. So it can provide references for the rational fertilization and the increase of calcium fertilizer utilization. 【Method】The experiment was carried out in a small shed under the condition of hydroponic culture and six calcium concentrations were designed. 【Result】The garlic seedling increments, pigment contents and photosynthetic parameters (Pn, E, Gs) of leaf increased with the increase of calcium concentrations in the range of 0-3.0 mmol•L-1, then they decreased with the increase of calcium concentrations ranging from 3.0 to 5.0 mmol•L-1. In addition, the SOD, POD, CAT, and NR activities of leaf and the root activity were enhanced at first and then receded with the increase of calcium concentrations, which were the strongest when the calcium was 3.0 mmol•L-1, but the MDA content in leaf showed a contrary tendency. Meanwhile, the fresh weight of bolt and bulb and the N, P, K, Mg contents in them got a peak value when the calcium concentration was 3.0 mmol•L-1, the Ca content in them was positively correlated to calcium concentrations. 【Conclusion】In conclusion, the 3.0 mmol•L-1 calcium concentration is the best treatment to garlic under hydroponic conditions.

Sequence Analysis and Prokaryotic Expression of Two Ethylene Responsive Factors MdAP2D4 and MdAP2D19

TIAN Yi-12, CHEN Ke-Qin-1, AN Xiu-Hong-1, HAO Hong-Mei-2, LIU Zhi-3, CONG Pei-Hua-2, HAO Yu-Jin-1

Scientia Agricultura Sinica. 2013, 46(17): 3635-3642. doi:10.3864/j.issn.0578-1752.2013.17.012

Abstract ( 572 )

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【Objective】 Cloning, sequence and expression analysis of two ERF (ethylene responsive factor) transcription factors from Malus domestica Borkh and expression of their fusion proteins in E. coli were determined to identify their disease resistant function and molecular mechanism.【Method】ESTs derived from suppression subtractive hybridization were blasted, and two EFR genes, MdAP2D4 and MdAP2D19, were cloned which contain these ESTs; Subsequently, phylogenetic relationship including the two apple proteins and Arabidopsis AP2 domain-containing proteins was analyzed using MEGA4.1 and the multiple alignment of AP2/ERF domain was constructed; Additionaly, the in vitro shoot cultures of ‘Gala’ apple were used to identify the two genes expression in response to exogenous MeJA; Finally, the two genes were inserted into vector pGEX-4T-1, and IPTG was used to induce the fusion proteins expression in E. coli BL21.【Result】The phylogenetic relationship of MdAP2D4 and MdAP2D19 was closer with Arabidopsis ERF proteins of group B3 than others. The transcript levels of MdAP2D4 and MdAP2D19 were higher in leaves than other organs and could be induced by exogenous MeJA. The results of SDS-PAGE demonstrated that the fusion proteins of the two genes could express in E.coli BL21 induced by IPTG.【Conclusion】 MdAP2D4 and MdAP2D19 are ERF transcription factors belonging to group B3, and they can be induced by exogenous MeJA.

Effects of Bud-Notching, Twisting Shoot and Removing Terminal Shoot on Branches Hydraulic Conductivity, Hormone Content and Flower Bud Formation of Fuji Apple

LI Xue-Wei-1, LI Bing-Zhi-1, LIU Fu-Ting-1, ZHANG Lin-Sen-1, HAN Ming-Yu-1, XU Sheng-Rong-2, ZHANG Yong-Wang-1, LIU Hang-Kong-1

Scientia Agricultura Sinica. 2013, 46(17): 3643-3650. doi:10.3864/j.issn.0578-1752.2013.17.013

Abstract ( 618 )

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【Objective】The aim of this study was to investigate the characteristics of Fuji branches hydraulic conductivity, hormone content and flower bud formation under the treatments of bud-notching, twisting shoot and removing terminal shoot, and then find the best method to promote flower bud formation.【Method】The 3-year-old field-cultivated Fuji (Malus domestica Borkh cv.Red Fuji /M26/M. sieversii (Ledeb) Roem) were used as materials to explore the changes of branches hydraulic conductivity, lateral buds hormone content, shooting percentage, flower bud formation and average yield per tree treated by bud-notching, twisting shoot and removing terminal shoot together after shoot-bending.【Result】Compared with the control, the integrated method of bud-notching, twisting shoot and removing terminal shoot together with shoot-bending significantly decreased stem and leaf area specific conductivity by 41.4% and 39.1%. The endogenous IAA, GAs in lateral buds treated with the integrated method decreased, while ZR, ABA content increased at flower bud differentiation stage. The shooting percentage enhanced by 32.7% in comparison with the control. The amount of blossoms and average yield per tree increased 2.3 times and 3.3 times more than those of control, respectively. The changes of branches hydraulic conductivity and lateral buds hormone content weakened tree vigor, while accelerated flower bud differentiation. The methods of bud-notching and removing terminal shoot after shoot-bending both had effects on Fuji flower bud formation, but the effects of the integrated method of bud-notching, twisting shoot and removing terminal shoot together with shoot-bending was most significant.【Conclusion】The integrated method of bud-notching, twisting shoot and removing terminal shoot together with shoot-bending inhibited tree vegetative growth, thus resulted in acceleration of flower development and increase of yield significantly.

Effect of Cavity Irrigation Underground on the Distribution of Soil Water Under the Canopy and Leaf Water Use Efficiency of Apple

BI Run-Xia, YANG Hong-Qiang, YANG Ping-Ping, FAN Wei-Guo, CHEN Jin-Pu, FAN Shu-Lei, WU Rui-Gang

Scientia Agricultura Sinica. 2013, 46(17): 3651-3658. doi:10.3864/j.issn.0578-1752.2013.17.014

Abstract ( 494 )

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【Objective】 The aim of this investigation was to elucidate the distribution of soil water under the canopy and leaf water use efficiency of apple by the cavity irrigation underground. 【Method】 Eight-year-old apple trees were used as materials in the experiment. The spatial and temporal distribution of soil water under canopy of trees and photosynthetic rate and water use efficiency of leaves were analyzed by comparing the cavity irrigation underground with storing water and fertilizer in the hollow and normal surface irrigation.【Result】The results showed that the soil water under canopy of trees rapidly declined after 3 days and it mainly concentrated in the range of 10-30 cm by the normal surface irrigation, and the area of water distribution presented a ‘plate shape’. The soil water rapidly declined after 7 days and it mainly concentrated in the 40-60 cm by the cavity irrigation underground. The soil water content had been maintained the highest level for a long time by cavity irrigation underground, and the area of water distribution that upside was less than the following liked ‘gourd shape’. The water was mainly distributed in the 10-60 cm deep soil zone similar to a ‘cylinder shape’ by the storing water and fertilizer in the hollow. Leaf chlorophyll content, photosynthetic rate and water use efficiency by cavity irrigation underground were the highest among the three irrigation models. 【Conclusion】 The soil water kept in the concentrated area of root and both photosynthetic rate and water use efficiency of apple leaf were improved by cavity irrigation underground.

Effect of Different Loading Methods on the Tobacco Quality and Safety During Flue-Curing

WU Sheng-Jiang-1, PAN Wen-Jie-1, GONG Chang-Rong-2, GENG Zhao-Liang-1, SUN Fu-Shan-3, LEI Bo-1, ZHAO Hui-Na-1, XIE Yi-Shu-1

Scientia Agricultura Sinica. 2013, 46(17): 3659-3668. doi:10.3864/j.issn.0578-1752.2013.17.015

Abstract ( 655 )

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【Objective】In order to discuss the formation mechanism of flue-cured tobacco quality and further improve tobacco quality and tobacco safety, the curing effect under different loading methods (T1: natural ventilation type curing; T2: hanging-pole bulk curing; T3: sticking leave bulk curing which is a kind of loose leaves bulk curing while tobacco leaves are stuck by skewer) was studied.【Method】Changes of moisture, carotenoids and cuticular components content of tobacco leaves during flue-curing, and the difference of the main chemical composition, neutral aroma components, smoke harmful ingredients content and sensory quality of flue-cured tobacco leaves under different loading methods were studied.【Result】The results showed that T3 had a higher moisture content than T2 at 48-72 h (P＜0.05). In terms of plastid pigment, T3 decreasing range of β-carotene and lutein content were 62.91% and 69.45%, higher than T1 (57.59% and 65.19%) and T2 (57.59% and 60.31%). T3 had a lower pigments content than T1 and T2 at the end of curing. In terms of cuticular components, T1 and T3 had a higher trichome exudates content than T2, and T1-T3 loss rate of trichome exudates were 8.82%, 54.05% and 10.52%, respectively. T3 had the highest alkanes content and its loss rate was 13.80% while T2 had the lowest alkanes content and its loss rate was 58.91%. Compared with T1 and T2, T3 had the highest neutral aroma components content (1 186.65 μg•g-1), a more coordinated chemical composition and higher score value (60.75 points) of sensory quality. In terms of smoke harmful ingredients content, the total carbonyl compound content of different treatments showed that T3 (1 034.08 μg•cig-1)＜T1 (1 144.76 μg•cig-1)＜T2 (1 251.19 μg•cig-1), and there was a significant difference between T1 and T2 (P＜0.05). The total smoke phenols content of tobacco leaves under different loading methods showed that T3 (149.93 μg•cig-1) ＜T2 (157.71 μg•cig-1)＜T1 (175.60 μg•cig-1). At the same time, T3 had the lowest content of crotonaldehyde, CO, and benzo[a]pyrene. Compared with T1, T2 and T3 had a lower content of HCN (P＜0.05).【Conclusion】Compared with T1 and T2, T3 had a slower dehydration speed of tobacco leaves during flue-curing, a lower carotenoid and smoke harmful ingredients content, and a higher cuticular components and neutral aroma components content, a more coordinated chemical composition and a higher score value of sensory quality of flue-cured tobacco leaves. Comprehensive evaluation of the flue-cured tobacco quality under different loading methods, T3 was the best.

Composition Change and Structure Identification of Phenols in Oat After Germination

FU Xiao-Yan-1, WU Qian-2, LI Shu-Yi-2, XIAO Juan-2, XIE Bi-Jun-2, SUN Zhi-Da-2

Scientia Agricultura Sinica. 2013, 46(17): 3669-3679. doi:10.3864/j.issn.0578-1752.2013.17.016

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【Objective】Changes of phenols composition after oat germination and structure identification of possible avenanthramides were studied. 【Method】 HPLC was used to analyze the phenol composition of extracts from ungerminated oat seeds, 6-day germinated oat grains, buds and roots. HPLC/Q-TOF-MS was chosen to detect the molecular weight of avenanthramides.【Result】The results indicated that the kind and content of phenolic acid in 6-day germinated oat grains were reduced. The relative content of phenolic acid was reduced from 32.14% to lower than 5%, while the relative content avenanthramide was increased from 45.52% to 79.21%. A group of avenanthramides such as 2c, 2p, 2f, t2, and t3 were both in ungerminated oat seeds and 6-day germinated oat grains. The molecular formula (molecular weight) of 2c, 2p, 2f, t2 and t3 were, respectively, C16H13O6N(315.0753), C16H13O5N(299.0818), C17H15O6N(329.0912), C18H15O5N(325.0968) and C19H17O6N(355.1077). Avenanthramides t2 and t3 increased drastically during oat germination. The composition of 6-day germinated oat buds and roots extracts was different from oat grain extract. The molecular weight of w6-w10 were, respectively, 564.1498, 592.1808, 494.1781, 710.1834 and 192.0433.【Conclusion】Phenol composition changed obviously after oat germination. The structure of six avenanthramides and the molecular weight of four possible avenanthramides were identified, thus offering a theoretical theory evidence for the study of germinated oat.

Effect of pH on Heat-Induced Gel of Myofibrillar Protein from Lamb M. longissimus dorsi Muscle

NI Na-12, WANG Zhen-Yu-1, HAN Zhi-Hui-3, HE Fan-1, PAN Han-1, MU Guo-Feng-1, ZHANG De-Quan-1

Scientia Agricultura Sinica. 2013, 46(17): 3680-3687. doi:10.3864/j.issn.0578-1752.2013.17.017

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【Objective】The effect of pH on myofibrillar protein heat-induced gel of M. longissimus dorsi sampled from Poll Dorset × Small Tail Han Sheep F1 lamb was studied, and the mechanism of pH influencing the formation of lamb myofibrillar protein heat-induced gel was also discussed.【Method】By measuring the water-holding capacity and hardness of lamb myofibrillar protein gels of different pH values, three typical pH values (pH 5.0, 6.0 and 7.5) were selected. Changes of molecular forces and thermal stability during heat-induced gelation were analyzed under the typical pH values. Difference between gel microstructures of three typical pH values was also compared.【Result】Myofibrillar protein heat-induced gels of lamb M. longissimus dorsi muscle showed different gel properties under three typical pH values. Protein gels had the lowest and a disorder microstructure at pH 5.0, and the lowest hardness was appeared at pH 6.0. The highest water-holding capacity and hardness were both appeared at pH 7.5, with a compact and ordered microstructure. The results of molecular forces analysis showed that hydrophobic interactions was the main forces during the gel formation, but pH influenced the ionic bonds and hydrogen bonds of the system. There might be some differences between the gel formation mechanisms under different pH values. 【Conclusion】 The overall results of this study demonstrated that molecular forces of heat induced gels were changed according to pH values, which resulted in different interactions of proteins or proteins and water, and then gels with different water-holding capacity, texture and microstructure were formed at different pH values.

Effect of h1-Calponin Gene on in Vitro Osteogenesis Differentiation of Bone Marrow Mesenchymal Stem Cells of Wuzhishan Minipig

HE Wei, WU Tian-Wen, HUANG Lei, ZHAO Wei-Min, XIA Ying, TANG Fang, MOU Yu-Lian, LI Kui

Scientia Agricultura Sinica. 2013, 46(17): 3688-3694. doi:10.3864/j.issn.0578-1752.2013.17.018

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【Objective】 The objective of the present study is to explore the relationship between h1-calponin gene and osteoblast differentiation of Wuzhishan minipig’s bone marrow mesenchymal stem cells (BM MSCs) and further understand the mechanism of osteoblast differentiation.【Method】BM MSCs were induced to osteoblast differentiation by adding osteogenesis differentiation medium. Alizarin Red staining was used to indentify the degree of osteoblast differentiation of BM MSCs. The expression profile of h1-calponin gene and the osteo-related genes osteopontin (OPN), osteocalcin (OCN), core binding factor α1 (Cbfα1) and core binding factor β (Cbfβ) were analyzed by quantitative PCR.【Result】Using Alizarin Red staining, it was found that the nodules of cells were increasing as the induction time went on. Q-PCR results showed that the expression patterns of OPN and OCN (two marker genes of osteogesis) were the same. Both OPN and OCN genes increased from day 0 to day 14, peaked at day 14 and finally declined. The osteo-related genes, Cbfα1 and Cbfβ, were highestly expressed at day 7 and then decreased gradually, but still higher than pre-induction. The expression of h1-calponin gene was gradually decreased during the induction and was nearly undetected from day 14 to 21.【Conclusion】Results of the experiment suggested there is a negative correlation between the expression of h1-calponin gene and osteoblast differentiation of Wuzhishan minipig’s BM MSCs is in vitro. The h1-calponin gene may be a negative-regulated gene during osteoblast differentiation.

Cloning of Xuhuai Goat SCD1 Gene, Subcellular Localization and the Preparation of Transgenic Mice

ZHU Cai-Ye, WEI Guang-Hui, LI Wei, WANG Dan, ZHENG Meng-Meng, LIU Zhi-Yong, ZHANG Ya-Ni, LI Bi-Chun

Scientia Agricultura Sinica. 2013, 46(17): 3695-3703. doi:10.3864/j.issn.0578-1752.2013.17.019

Abstract ( 523 )

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【Objective】The aim of this paper was to clone the stearoyl-CoA desaturase 1(SCD1) gene from Xuhuai goat, and analyze it’s sub-cellular localization through enhanced green fluorescent (EGFP) fusion protein. Study the gene expression in heterologous animals, explore the possibility of preparing dissimilar transgenic animals and verify whether the foreign gene can stably inherited. 【Method】cDNA was cloned by reverse transcription polymerase chain reaction (RT-PCR), it’s biological information characteristics was analyzed by online software, and fusion expression vector named pEGFP-C1-SCD1 was constructed successfully. Then NIH-3T3 cells were transfected with pEGFP-C1-SCD1 through lipofectamine TMLTX&PLUS(LTX). At 48 h post-transfection, the transfected cells were observed under fluorescence inverted microscope. Then the mRNA expression level of SCD1 was detected by RT-PCR. The transgenic mice was produced by testicular injection, and the target gene expression in the F1 and F2 generations of individual was detected at DNA and protein levels. 【Result】 The results showed that the coding sequence (CDS) length of Xuhuai goat SCD1 gene was 1 074 bp, encoding 357 amino acids with GenBank accession number (JX854036). The SCD1 cDNA coding sequence was compared with the corresponding region of Human, Rattus norvegicus, Mus, Ovis aries, Bos taurus, Sus scrofa, respectively, the similarity was 86%, 83%, 82%, 98%, 95%, and 90%, respectively, and amino acid sequence homology was 84%, 79%, 79%, 96%, 92%, and 95%, respectively. pEGFP-SCD1 was successfully constructed. The RT-PCR results showed the SCD1 mRNA expressed successfully in vitro. The SCD1 protein was localized in the cytoplasm which was in line with the result of online prediction. The gene can also be expressed in mice transiently and persistencely after intravenous and testicular injection and the gene can be genetically stable. 【Conclusion】The SCD1 gene cDNA of Xuhuai goat was cloned successfully and in the single-cell level can express in the cytoplasm, also can express in mice.

Polymorphisms of Chicken ApoA5 Gene and Their Associations with Cholesterol Content and Reproductive Traits

DU Xiao-Hui, XIONG Ting, LIU Jun-Ying, ZHANG Huai-Yong, WAN Huo-Fu, ZHU Qing

Scientia Agricultura Sinica. 2013, 46(17): 3704-3711. doi:10.3864/j.issn.0578-1752.2013.17.020

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【Objective】To balance the relationship between cholesterol concentration and reproductive traits, the associations among the traits of cholesterol and reproduction and polymorphisms of chicken ApoA5 gene were studied. 【Method】 The reproduction traits were recorded and the cholesterol concentrations of serum (SCC) and egg yolk (YCC) were measured in 434 SD hens at 300th day. Through PCR-SSCP and PCR-RFLP analyses, 6 new SNPs were detected in ApoA5 gene and their associations with the traits of cholesterol and reproduction were analyzed. 【Result】The effects of the single SNP mainly focused on the age of first egg (AFE) and on the SCC. The results of association analysis of the haplotype, constructed with SNP2, SNP4 and SNP5 because of their closer location and consistent effects, were roughly accordant with the single SNP, but the trait differences between haplotype combinations were enlarged. The haplotype combinations (H5H8, H4H8, H5H6, H6H6) with higher SCC and YCC had better reproductive performance, and H1H5, H1H6 and H3H8 with lower SCC and YCC were on the contrary.【Conclusion】Haplotype SNP2-4-5 of ApoA5 gene revealed the differences of cholesterol and reproductive traits among various combinations, and could be used as a molecular markers to improve reproductive traits.

The Profiles of Related Genes mRNA Expression in Duck Hypothalamus-Pituitary Growth Axis During Early Development

HU Yan, XU Wen-Juan, LIU Hong-Xiang, SONG Wei-Tao, SONG Chi, TAO Zhi-Yun, DAN Yan-Ju, LI Hui-Fang

Scientia Agricultura Sinica. 2013, 46(17): 3712-3720. doi:10.3864/j.issn.0578-1752.2013.17.021

Abstract ( 551 )

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【Objective】The present experiment was conducted to study the tissue expression characteristics of related genes mRNA expression in duck hypothalamus-pituitary growth axis during embryogenesis and post-hatching development in different breeds of ducks. 【Method】We have compared the ontogeny of body weight and expression of growth hormone release hormone (GHRH), somatostatin (SS), growth hormone (GH), growth hormone receptor (GHR), insulin-like growth factor-I (IGF-I) mRNA on the embryonic day 13, 17, 21, 25, and 27 and the 7-day-old Gaoyou duck and Jinding duck. 【Result】At the first time, the trial presented that duck hypothalamic GHRH and SS, pituitary GH, liver GHR and IGF- I mRNA were detected at E13. Except SS maintained a lower expression status in the duck early development, the other four genes showed a extremely significant difference in variety and time. In addition to GH, duck GHRH, GHR and IGF-I mRNA expression were significantly affected by breed and age interaction during early development. Compared with previous studies on chicken, significant species differences were observed in poultry embryonic hypothalamus-pituitary growth axis related gene mRNA expression. During early development, extremely significant variety and time specificity was found in developmental change of body and liver weight, and varying degrees of linear correlation was showed between that and related genes mRNA expression in duck hypothalamus-pituitary growth axis. 【Conclusion】It was suggested that different genetic backgrounds may lead to the differential growth rate and the related gene differential expression, and the differential expression of related genes mRNA in duck hypothalamus-pituitary growth axis may be involved in the early growth of ducks.

Effects of Mating Flight on sRNAs Expression in Sexual Matured Virgin Queens (Apis cerana cerana)

WU Xiao-Bo, WANG Zi-Long, SHI Yuan-Yuan, ZHANG Fei, ZENG Zhi-Jiang

Scientia Agricultura Sinica. 2013, 46(17): 3721-3728. doi:10.3864/j.issn.0578-1752.2013.17.022

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【Objective】The objective of this study is to analyze the influence of flight behaviour on sRNAs expression in matured virgin queens (Apis cerana cerana). 【Method】The sexual matured virgin queens of A. cerana cerana were used as flying group that allow to fly in a certain area and the other group was not. The sRNAs expression differences between the two groups were analyzed by using a high-throughput sequencing method. 【Result】Both of the flying matured virgin queens and non-flying counterparts had a complicated sRNAs population, and that the length of sRNAs varied, 22 nt and 27-29 nt being the predominant length, while the frequence percent of the same RNA was different in the two samples. Combining deep sequencing and bioinformatics analysis, the percent of the common sequences in total sRNAs was 92.79%, but the number of unique sRNAs in the flying queens was more than that in non-flying queens. Matching to known miRNA alignment, a total of 25 known miRNAs were significantly different between them, with 1 up-regulated and 24 down-regulated in the flying matured virgin queens while there were 11 target genes of 19 known miRNA which were expressed differentially between them.【Conclusion】There are a large number of sRNAs changes during the flight process of matured virgin queens which may play an important role in adjusting physiology for mating successfully of virgin queens.

Exploration and Application of Non-Denaturing FISH on Chromosomes Research in Sesame (Sesamum indicum L.)

LIU Yan-Yang, CUI Cheng-Qi, MEI Hong-Xian, WU Ke, ZHENG Yong-Zhan

Scientia Agricultura Sinica. 2013, 46(17): 3729-3735. doi:10.3864/j.issn.0578-1752.2013.17.023

Abstract ( 560 )

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【Objective】The objective of this paper is to study the optimal experimental conditions for non-denaturing FISH (ND-FISH), and to explore its application for chromosome recognition in sesame. 【Method】 Using oligomeric nucleotide (AC)8 as probe, the effects of different elution strengths, digestions and hybriding times on the ND-FISH were analyzed to optimize the protocol. 【Result】The optimal experimental conditions for ND-FISH were as follows: cells on slides were treated with DNase-free RNase A at a concentration of 100 μg•mL-1 for 1 h, 6 µL of the probe (20 ng•µL-1), hybridization for 4 h, elution with the 4×SSC/0.2% Tween 20 optimal solution for 10 min. Using (AC)8 as a probe, mitotic metaphase chromosomes of Yuzhi 11 were analyzed by ND-FISH. Nine pairs of the chromosomes of Yuzhi 11(seven pairs of chromosomes had strong signals, and two pairs of chromosomes had weak signals) could be distinguished based on the hybridization signals combining with the length and arm ratio of chromosomes. 【Conclusion】ND-FISH technology with (AC)8 as a probe was used to successfully identify 18 of 26 chromosomes of cultivated sesame for the first time.

Effects of Hydroponics on Nutrient Components and Nitrate Contents in Tips of Leaf Vegetable Sweet Potato

CHEN Xuan-Yang, ZHANG Zhao-Juan, ZHENG Jia-Wei, LIN Yu-Li

Scientia Agricultura Sinica. 2013, 46(17): 3736-3742. doi:10.3864/j.issn.0578-1752.2013.17.024

Abstract ( 626 )

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【Objective】The effects of hydroponics with different nutrient solutions, nutrient solutions with different N forms and their ratio on nutritional quality and nitrate content in tender stem, leaf and petiole of the tips of leaf vegetable sweet potato were studied. 【Method】 Half-Hoagland(1/2HL) nutrient solution was used for hydro culturing leaf-vegetable sweet potato, the leaves, stems and petioles were sampled for determining the contents of nutrient components and nitrate after culturing for 3, 5 and 7 weeks. Four classical selected nutrient solutions and 10 other nutrient solutions modified with different N forms and ratios were used, nitrate content of leaf, stem and petiole of samples were determined after hydro cultured for 5 weeks. 【Result】The contents of VC, flavone and nitrate in leaf-vegetable sweet potato hydro cultured in the 1/2HL nutrient solution were significantly higher than that of cultured in the field, while there were no difference in protein contents between them. There were significant differences in nitrate content among different parts of leaf vegetable sweet potato plant, the content in the tender stem was the highest, the leaf was the medium, and the petiole was the lowest. There were significant effects of different nutrient solutions on the nitrate content in the leaf-vegetable sweet potato. The nitrate content in leaf-vegetable sweet potato cultured in Hua’nan B and 1/2HL solution were significantly higher than that in Hua’nan A. While the nitrate content in Yuansi solution was similar with that cultured in the field, and both of them were significantly lower than those in 1/2HL, Hua’nan A and Hua’nan B. There were significant effects of different N forms and their ratios in the nutrient solutions on the nitrate content in leaf-vegetable sweet potato. With the increasing of concentration and proportion of NO3--N in the solution, the nitrate content in leaf vegetable sweet potato increased. The appropriate content of NH4+-N in the solution retarded the increase of nitrate content in the leaf-vegetable sweet potato. (CO(NH2)2-N in the nutrient decreased the nitrate content of hydroponic leaf-vegetable sweet potato. 【Conclusion】Hydro culturing increased VC, flavone and nitrate contents in leaf-vegetable sweet potato, while had no influence on the protein content. More NO3--N in the nutrient solution increased the nitrate content of the leaf-vegetable sweet potato, while appropriate content of NH4+-N retarded its increase. (CO(NH2)2-N in the solution could decrease the nitrate content.

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