Scientia Agricultura Sinica

Comparison of Panicle Length QTL Based on F2 and F8 Populations Derived from Rice Subspecies Cross

GUO Xiao-Jiao-2, ZHANG Tao-13, JIANG Kai-Feng-13, YANG Li-13, CAO Ying-Jiang-1, YANG Qian-Hua-1, YOU Shu-Mei-1, WAN Xian-Qi-3, LUO Jing-1, LI Zhao-Xiang-1, GAO Lei-1, ZHENG Jia-Kui-123

Scientia Agricultura Sinica. 2013, 46(23): 4849-4857. doi:10.3864/j.issn.0578-1752.2013.23.001

Abstract ( 486 )

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【Objective】In this study, rice panicle length QTL was analyzed by using F8 RIL population, the similarities and differences of the genetic map, QTL locations, QTL effects in different derivative population of the same parent were compared and the stable expression of QTL of panicle length trait was identified. The results of the study will provide more information about the genetic behavior of rice panicle length and be benefit to the MAS (marker-assisted selection).【Method】A F8 population containing 188 lines, which was derived from the cross between the indica variety Luhui99 and the japonica variety Nipponbare, was analyzed with a genetic linkage map consisting 207 DNA markers. The statistic software of QTL Network 2.0 was applied to detect QTL and QTL effect analysis for rice panicle length. The similarities and differences of QTL mapping and genetic map between F8 and F2 population were discussed. 【Result】A total of 7 QTL with significant additive effects were detected on chromosomes 2, 3, 6, 7, 8 and 10 in F8 population. The proportions of phenotypic variance explained by individual QTL ranged from 3.38% to 14.8% and the QTL could jointly explain 52.5% of the total phenotypic variance. There were five same chromosomes having rice panicle length QTL in F8 and F2 genetic map and most of the QTL’s physical locations are overlapping or containment. The F8 and F2 genetic maps differed in molecular marker number, marker arrangement, genetic distance and average distance between the markers. 【Conclusion】 Seven rice panicle length QTL were detected in F8 RIL population. Although there were many differences betwen F8 and F2 populations in genetic map, QTL analysis and so on, a steady main QTL was found on the sixth chromosome. In addition, four stable rice panicle length QTL were detected in the study which were not reported in previous references.

Development and Characterization of Both PvPGIP2 and TaLTP5 Transgenic Wheat Lines with Resistance to Take-all

RONG Wei-12, WANG Jin-Feng-2, LI Zhao-2, WANG Ai-Yun-1, DU Li-Pu-2, YE Xing-Guo-2, WEI Xue-Ning-2, ZHANG Zeng-Yan-2

Scientia Agricultura Sinica. 2013, 46(23): 4858-4867. doi:10.3864/j.issn.0578-1752.2013.23.002

Abstract ( 349 )

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【Objective】 Take-all is a destructive disease of wheat production worldwide. The aim of this study is to develop and select stable PvPGIP2 and TaLTP5 transgenic wheat lines with resistance to take-all pathogen Gaeumannomyces graminis. 【Method】Gene recombination technology was used to prepare the transformation vector pA25-PvPGIP2-TaLTP5 expressing both PvPGIP2 and TaLTP5. Particle bombardment method was used to introduce both PvPGIP2 and TaLTP5 into wheat cultivar Yangmai 18. PCR, RT-PCR and qRT-PCR methods were used to detect the presence and transcript levels of PvPGIP2 and TaLTP5 in the transgenic wheat plants of T0-T4 generations. G. graminis mycelia plug inoculation and take-all severity and the disease index were used to score the resistance degrees of these transgenic wheat lines and non-transgenic wheat Yangmai 18 at young seedling-stage. 【Result】The results indicated that the introduced PvPGIP2 and TaLTP5 genes were stably inherited, and could be highly expressed in six transgenic wheat lines. The transgenic wheat plants expressing PvPGIP2 and TaLTP5 showed significantly enhanced resistance to G. graminis compared with non-transgenic wheat Yangmai 18. 【Conclusion】 These results suggested that the introduced PvPGIP2 and TaLTP5 genes can be used for improving wheat resistance to take-all.

Study on Flora of National Key Preserved Agricultural Wild Plants in North China Region

YANG Xiang-Fu, SHI Xue-Zhen, LI Jing-Yuan, WANG Tai-Xia

Scientia Agricultura Sinica. 2013, 46(23): 4868-4875. doi:10.3864/j.issn.0578-1752.2013.23.003

Abstract ( 462 )

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【Objective】This article aims to analyze the elements of national key preserved agricultural wild plants in North China region, and to provide reference for protection and sustainable utilization of plant resources.【Method】On the basis of the geographical distribution of modern pteridophytes in China, the areal-types of seed plants and D. Szymkiewicz’s similarity coefficient in genera were analyzed. 【Result】 There are 44 species of national key preserved agricultural wild plants in North China, belonging to 39 genera and 24 families. They are mainly temperate in nature. At the generic level, various temperate genera, tropical genera and endemic genera to China contributed 72.22%, 13.89% and 13.89% of the total (excluding cosmopolitan genera). At the species level, various temperate elements, tropical elements and endemic to China contributed 54.76%, 2.38% and 42.86% of the total species. Compared with the flora of two neighboring areas, the flora is much more closely allied to northeast China than to northwest China.【Conclusion】There are 7 areal-types and 2 subtypes of families, 12 areal-types and 6 subtypes of genera and 6 areal-types and 3 subtypes of species in the geographical distribution.

Effects of Different Types of Bowl Seedlings and Densities on Characteristics of Nitrogen Uptake, Utilization and Translocation of Bowl Transplanted Japonica Super Rice

XU Ke-1, ZHOU Xing-Tao-1, 2 , CAO Li-Qiang-1, ZHANG Hong-Cheng-1, GUO Bao-Wei-1, CHEN Hou-Cun-3, WU Zhong-Hua-3, ZHU Cong-Cong-1, YANG Yan-1

Scientia Agricultura Sinica. 2013, 46(23): 4876-4892. doi:10.3864/j.issn.0578-1752.2013.23.004

Abstract ( 391 )

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【Objective】 The objective of this experiment was to identify the differences in N absorption and use efficiency of three different bowl types and densities in bowl transplanted rice. 【Method】 The experiment was conducted by the seedlings of 3-bowl combined together, 2-bowl combined together and single bowl seedling disc with Wuyunjing 24 and Nanjing 44 as materials, and the setting of different transplanting densities. The differences in N absorption, use efficiency and their relationship with yield were studied. 【Result】 For the same type of bowl seedlings, N uptake amount at heading and maturity stages, recovery efficiency, agronomic efficiency, partial factor productivity increased first and then decreased with the increase of the basic seedlings number. Under the same basic seedling conditions, the N uptake amount at heading and maturity stages, recovery efficiency, agronomic efficiency and partial factor productivity showed the following rules: the lower basic seedling number condition of 36×104•hm-2 to 54×104•hm-2 showed a trend of single bowl＞2-bowl＞3-bowl, the medium basic seedling number condition of 72×104•hm-2 showed a trend of 2-bowl＞single bowl＞3-bowl, while the larger basic seedling number condition of 90×104•hm-2 to 108×104•hm-2 showed a trend of 2-bowl and 3-bowl＞single bowl. Under the three maximum yield conditions, the N uptake at heading and maturity stages, recovery efficiency, agronomic efficiency and partial factor productivity showed a trend of 2-bowl＞3-bowl＞single bowl. Correlation analysis of the three maximum yield treatments indicated that there existed a significant or a highly significant positive correlation between yield and N uptake of leaves and aboveground at maturity stage, N uptake from jointing to heading stage and heading to maturity stage.【Conclusion】Under the suitable or larger basic seedling number conditions, compared with single bowl cultivation, 3-bowl and 2-bowl, especially the 2-bowl seedling planting methods have a higher N absorption amount at middle and late growth stages and N use efficiency is also high. They are new ways with high nitrogen use efficiency, labor saving to accomplish super high yielding cultivation.

Water Consumption Characteristics of Winter Wheat with Soil-Coated Ultrathin Plastic-Film Mulching Under Different Water Managements

LI Meng-Zhe-1, ZHANG Wei-Hong-2, ZHANG Yong-Sheng-1, DANG Hong-Kai-3, WANG Lei-1, HE Li-Qian-1, LI Ke-Jiang-3, LI Yan-Ming-1, DU Xiong-1

Scientia Agricultura Sinica. 2013, 46(23): 4893-4904. doi:10.3864/j.issn.0578-1752.2013.23.005

Abstract ( 443 )

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【Objective】In the light of the actual conflict between severe deficiency of water resource and more water consumption of winter wheat production in Haihe Plain, a field experiment was conducted to study the water consumption characteristics of winter wheat planted with soil-coated ultrathin plastic film mulched under different water managements, and the objective of this study was to search out a new water saving method for winter wheat production.【Method】The test lasted for three winter wheat growing years in two sites, and a field experiment was adopted with a conventional wheat planting method as a control.【Result】Under soil-coated ultrathin plastic film mulching, the field water consumption of winter wheat was significantly decreased by 116.1-167.9 mm in a whole growing season, and the soil water consumption accounted for 55.5%-77.9% of the total. With plastic film mulching, the water consumption before wheat recovery was reduced significantly so that the water consumption percentage from jointing to harvest was increased. Under soil-coated plastic film mulching condition, the water intake depth in soil body was 40 cm shallower than that under bare field, the 2 m-deep soil layer could supply 212.2-240.3 mm water for wheat in the whole growing season. As one-time 75 mm irrigation in spring was practised, the winter wheat uptook the water from 0-140 cm soil layer, which was 40 cm deeper than the control that was irrigated with the total 225 mm water for three times, soil water consumption was decreased significantly in 2 m-deep layer as the one-time 75 mm irrigation delayed. Under soil-coated ultrathin plastic film mulching condition, the rain-fed wheat after sowing achieved a yield more than 7 500 kg•hm-2 with a 284.3 mm filed water consumption, the wheat irrigated with over 60 mm water before sowing or 75 mm water at heading stage could achieve an equivalent yield to the control, and the water use efficiency was increased by more than 40% compared with the control, the net water use efficiency and irrigation water use efficiency were the best among all the treatments.【Conclusion】Soil-coated plastic film mulching and rain fed or with little irrigation timely is a new way for winter wheat production as it can considerably decrease water consumption and relieve the intense conflict between water resource supply and requirement in Haihe Plain.

Analyzing the Influence of Drip Irrigation Uniformity on Spring Maize Yield Using the Spatial Effect Model

WANG Zhen, LI Jiu-Sheng, ZHANG Hang, LI Yan-Feng

Scientia Agricultura Sinica. 2013, 46(23): 4905-4915. doi:10.3864/j.issn.0578-1752.2013.23.006

Abstract ( 371 )

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【Objective】The objectives of this study were to evaluate the influence of drip system uniformity and soil spatial variability on the spring maize yield. 【Method】Field experiments were conducted during four growing seasons of spring maize (Zea may L.) from 2009 to 2012 in the North China Plain. Three Christiansen uniformity coefficients (Cu) of 0.66, 0.81, and 0.99 and three levels of irrigation amount (50%, 75%, and 100% of irrigation requirement) were used in 2009 and 2010. While three Cu of 0.59, 0.80 and 0.97 and three levels of nitrogen applied at 0, 120 and 210 kg•hm-2 were used in 2011 and 2012. The influence of drip irrigation uniformity on spring maize yield was analyzed using a spatial effect model and an analysis of variance (ANOVA) approach. 【Result】The ANOVA results indicated that neither drip uniformity nor irrigation amount and nitrogen application level imposed a significant influence on maize yield. However, the spatial effect model, which considered the influence of spatial variability in the field on spring maize yield, demonstrated that nonuniformly applied water and fertilizers may imposed a negative influence on growth of crop when an extremely low uniformity (e.g. Cu＜0.6) was used. 【Conclusion】This study recommended that the spatial variability of field should be considered when the target drip irrigation uniformity is determined. For the semi-humid regions such as the North China Plain, an extremely low drop uniformity of less than 0.6 is not recommended.

Identification and Pharmaceutical Screening of Brown Spot Disease on Actinidia chinensis

ZHAO Jin-Mei-1, GAO Gui-Tian-1, GU Liu-Jie-1, SUN Xiang-Yu-1, XUE Min-1, GENG Peng-Fei-1, LEI Yu-Shan-2

Scientia Agricultura Sinica. 2013, 46(23): 4916-4925. doi:10.3864/j.issn.0578-1752.2013.23.007

Abstract ( 473 )

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【Objective】The objective of this study is to identify the pathogenic fungus of a serious disease which had caused brown spot disease to Actinidia chinensis, and screen out effective chemicals for inhibition of its mycelium growth.【Method】 Isolates from diseased fruits and leaves were cultured on PDA medium and purified by single spore culture, respectively. Identification of the pathogenic fungus was carried out according to the characteristics of fungal morphology, pathogenicity and by the molecule biology identification. Eighteen fungicides were used to determine the bacteriostatic effect on that pathogen, and the toxicity regression equation of them was also established.【Result】Fungal morphological characteristics of three purified pathogenic strains from different samples were the same, and then these strains were identified to the same pathogen, Alternaria alternata (Fr.:Fr.) Keissler, by rDNA-ITS sequence alignment and BLASTn tool combined with morphology characteristics. On 10 μg•mL-1 drug-containing PDA plates, the average inhibition rate of the pathogen was more than 80% by seven kinds of fungicides which were 400 g•L-1 flusilazole, 500 g•L-1 iprodione, 80% tebuconazole, 10% multi-antimycin B, 12.5% diniconazole, 33.5% quinolinone and 10% difenoconazole, respectively.【Conclusion】It is concluded that A. chinensis brown spot disease is caused by A. alternata, and 400 g•L-1 flusilazole, 500 g•L-1 iprodione, 10% multi-antimycin B and 10% difenoconazole have a better inhibitory effect on A. alternata than others used in this research.

Rapid PCR Detection Technology of Common Scab Pathogens in China

GUO Feng-Liu, ZHANG Hai-Ying, YU Xiu-Mei, ZHAO Wei-Quan, LIU Da-Qun

Scientia Agricultura Sinica. 2013, 46(23): 4926-4932. doi:10.3864/j.issn.0578-1752.2013.23.008

Abstract ( 425 )

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【Objective】The composition diversity of common scab pathogens was confirmed by previous researchers. A qualitative detection method for potato scab pathogens was established in this work by screening the universal probe primers among different pathogens. 【Method】 Four typical pathogenic strains which belonged to Streptomyces scabies (CPS-1), S. galilaeus (CPS-2), S. acidiscabies (CPS-3) and S. turgidiscabies (CPS-4) were selected to extract genome DNA. The primers were designed according to the sequences of four genes txtA, txtB, txtC (P450) and txtD (nos) that were from pathogen-specific toxin biosynthesis gene cluster what had registered in the NCBI GenBank. All primers were used for amplification for screening a high specificity and stability primers. After temperature gradient filter for optimizing the reaction system, a detection technology of potato scab pathogens was established. 【Result】 The universal primers B1/B2 obtained in this work showed a good specificity to all tested pathogenic strains. The stable bands were amplified among the pathogenic strains, but nonpathogenic strains didn’t have that band. By using the spore dilution method, the sensitivity of PCR detection method was determined to 20 pg•μL-1. The detection value of the threshold was 4.0 CFU/μL for pathogen spores. The PCR detection results showed that the target bars could be observed in pathogenic Streptomyces, scabby tissue and diseased field soil samples. While the nonpathogenic Streptomyces, healthy potato tissues, scab-free field soil and other test strains did not have corresponding objective band. That suggested the PCR detection method established in this work was specific and stable to common scab pathogens. 【Conclusion】 A qualitative detection method for potato scab pathogens was established in this study, which can quickly detect pathogenic strains from diseased tissues and soil samples.

Action of the Metabolite Produced by Streptomyces cacaoi Strain 182-2 on Alternaria alternata

GAO Fen-12, WU Yuan-Hua-2, WANG Meng-Liang-1

Scientia Agricultura Sinica. 2013, 46(23): 4933-4940. doi:10.3864/j.issn.0578-1752.2013.23.009

Abstract ( 432 )

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【Objective】 The objective of this study is to understand the antifungal activity and mechanism of KA08 produced by Streptomyces cacaoi strain 182-2 against Alternaria alternata so as to provide a scientific basis for further product development and application.【Method】Estimation of mycelia wet weight and spore germination in petri dish method were used to test the inhibitory effect of KA08 on mycelia growth and spore germination. Electrical conductivity method was used to test the permeability of pathogen plasma membrane, and the ultraviolet absorption method was employed to detect the content of ergosterol, MDA and soluble protein.【Result】KA08 had a remarkable inhibition effect on mycelia growth and spore germination. It caused shorter and twisted nodes, abnormal tubes with tips expanded or deformed, or saclike expansions. Twenty-four hours after treatment, the EC50 of KA08 against conidial germination of A. alternata was 177.53 µg•mL-1. Studies on the action mechanism of KA08 found out that the electrical conductivity of the cultural filtrate increased obviously after the pathogen was treated with KA08, indicating the leakage of protoplasm from the mycelia. The ergosterol content in the cell membrane of mycelia decreased, remarkably. The content of MDA in the mycelia and the cultural filtrate increased notably, and the protein content in the mycelia reduced greatly.【Conclusion】KA08, the active metabolite of strain 182-2, has obvious inhibitory activities against A. alternata. Its bioactivity lies in its doing damage to the pathogen cell membrane and increasing the permeability of plasma membrane through inhibiting the synthesis of ergosterol in the cell membrane and causing lipid peroxidation of the cell membrane, which resulted in the poor and abnormal mycelia growth. These results suggest that cell membrane is one of the main action sites of KA08.

Adaptability and Physiological Response to Host Plant Species Switching in Apolygus lucorum

XIAO Liu-Bin, TAN Yong-An, SUN Yang, ZHAO Hong-Xia, WU Guo-Qiang, BAI Li-Xin

Scientia Agricultura Sinica. 2013, 46(23): 4941-4949. doi:10.3864/j.issn.0578-1752.2013.23.010

Abstract ( 410 )

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【Objective】The objective of this study is to clarify the effects of plant species switching on reproduction, growth and physiological responses of the experimental population of green blind bugs (Apolygus Lucorum).【Method】The life tables of A. lucorum in a laboratory population were constructed to systematicly study the effect of different species hosts (Phaseolus vulgaris, Vigna unguiculata, V. radiata, Sumian9, Ezamian10). The activities of trehalase, amylase and proteinase and the content of trehalase were examined in laboratory populations of A. lucorum that reared on different species of host plants.【Result】The developmental periods, survival rates, body weight of 5th instar, longevity and hatching rate of A. lucorum were significantly different among the five species host plants. Population increased from high to low was P. vulgaris (16.56)＞Ezamian10 (3.23) ＞Sumian9 (2.14) ＞V. unguiculata (1.67) ＞V. radiate (1.31) by comprehensive analysis of life tables. At the same time, there was a certain difference of the trehalase activity and the content of trehalose of A. lucorum on different host plants. The trehalase activity for Ezamian 10 came to the peak was at 3.74 μg•mg-1•min-1 (P＜0.05), however, the trehalose content came to the valley was at 3.03 μg/adult (P＜0.01).【Conclusion】As green blind bugs on P. vulgaris transferred to the other four species host plants, life parameters decreased, such as population trend index and so on. The frequency differences were obviously among these five populations. In general, A. lucorum has high adaptability to cotton.

3D Reconstruction and Porosity Computing of Soil Based on SFS Algorithm

WANG Xiao, WEN Wei-Liang, GUO Xin-Yu, WANG Chuan-Yu

Scientia Agricultura Sinica. 2013, 46(23): 4950-4956. doi:10.3864/j.issn.0578-1752.2013.23.011

Abstract ( 337 )

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【Objective】In order to calculate the characteristics of soil structure and porosity quickly and accurately, a method based on SFS algorithm (shape from shading) is proposed for 3D reconstruction of soil. 【Method】This method deduced the illumination equation based on Lambertian surface reflection model, and then solved this equation for working out the pixels’ height of soil’s gray microscope images with the method of Taylor expansion and Jacobi iteration. 【Result】Experimental results demonstrate that compared with the soil porosity measured by soil sampler and baking, this method has a result with 0.81% less and 1.75% error rate. 【Conclusion】The proposed method, by using soil microscope images’ 3D information, could calculate the soil physical characteristics like porosity more objectively and accurately.

Effects of Different Irrigation and Cropping Years on Waxy Corn Growth and Yield Using Drip Irrigation with Plastic-Film Mulching on Saline-Sodic Soils in Arid Area

TAN Jun-Li-13, KANG Yue-Hu-2, DOU Chao-Yin-4

Scientia Agricultura Sinica. 2013, 46(23): 4957-4967. doi:10.3864/j.issn.0578-1752.2013.23.012

Abstract ( 357 )

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【Objective】Soil environment had changed greatly during the reclamation of saline-sodic soils with drip irrigation technology. Crop growth is the comprehensively reflection of this change. In this paper, the response of waxy corn growth and yield to changes of soil environment during the reclamation process of saline-sodic soils was investigated.【Method】Through the method of field experiment, the emergency rate, height, the stem perimeter, the dry biomass, the yield of waxy corn and irrigation water use efficiency in the fields of reclaimed and drip irrigated for 1 year, 2 years, 3 years and 4 years were determined and the response mechanism of waxy corn to soil environment changes was investigated. 【Result】Results showed that using the irrigation method that irrigated about 30 mm water at first time after crop sowing and then used tensiometer, which was buried at 0.2 m depth immediately under drip emitter, to control soil matric potential higher than -10 kPa could guarantee emergence rate above 90% and high yield of waxy corn. Even in the field with drip-irrigation and cropped in the first year.The height, the stem perimeter, the dry biomass, the yield of waxy corn and irrigation water use efficiency increased with the increase of irrigation and cropping years. There was a significant difference in these indexes among different irrigation and cropping years during the corn growth period. In addition, the growth rhythm of waxy corn in different treatments had significant differences also. 【Conclusion】Application of drip irrigation and cropping technology introduced in this paper for reclaimation of saline-sodic soils, soil environments changed greatly with the increase of the irrigation and cropping years. The changes of saline-sodic soils were reflected by the growth index and yield of waxy corn.

Screening and Evaluation of Germplasms with High Resistance to Fruit Cracking in Ziziphus jujuba Mill.

YUAN Zan-1, LU Yan-Qing-1, ZHAO Jin-2, LIU Meng-Jun-1

Scientia Agricultura Sinica. 2013, 46(23): 4968-4976. doi:10.3864/j.issn.0578-1752.2013.23.013

Abstract ( 452 )

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【Objective】Fruit cracking is the most serious physiological disease in the production of Chinese jujube, the objective of this study was to screen and evaluate the germplasms with high resistance to cracking. These germplasms would provide materials for planting and breeding of Chinese jujube directly.【Method】The cracking resistance of 169 cultivars were evaluated for 5 years by comparison of cracking type, cracking rate and degree of cracking extent. Then, the fruit microstructures of cultivars with different degrees of cracking resistance were compared by using paraffin slice. 【Result】The grading system of jujube fruit cracking was established according to the length and area of fruit cracking. Statistical analysis of four fruit cracking types showed that longitudinal crack was the main type. Some resistant cultivars, such as Huluchangzao, Guantanzao, Hebeilongxuzao, Chahuzao, Xuezao, Jianzao, Chuanlingzao, Chengwudongzao, Zhongyangtuanzao, Hongzhaocuizao, Mianxuzao, Lengzao, Shaizao, Wanniunaizao, Nanjingdamuzao and Mopanzao, were screened out. The fruit structures of cultivars with different cracking resistance degrees were significantly different. The peel epidermis layer of resistance cultivar is of uniform thickness, with cells closely arranged in neat rows. The weaker the resistance to fruit cracking, the unevener of the peel epidermis layer, and the more unordered the cells. 【Conclusion】 Several cultivars with high resistance to fruit cracking were found. The resistance to fruit cracking is positively correlated to the density and uniformity of fruit epidermal cells but not the density of pulp cells.

Diversity of Resistance to Jujube Witches’ Broom Disease Among the Strains of Ziziphus jujuba Mill. Junzao

XIAO Jing-1, YANG Yan-Rong-1, ZHAO Jin-2, LIU Meng-Jun-1

Scientia Agricultura Sinica. 2013, 46(23): 4977-4984. doi:10.3864/j.issn.0578-1752.2013.23.014

Abstract ( 421 )

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【Objective】Sixteen strains of Z. jujuba Junzao were collected and evaluated, aiming at revealing the diversity of resistance to JWB within cultivar, screening excellent strains with high resistance. 【Method】The JWB symptom development and phytoplasma infection were studied through field investigation and PCR analysis. The fruit economical characters, including the rate of water content, fruit weight, fruit shape index, ratio of edibility, sugar content, Vc content and titratable acidity, were comprehensively evaluated among different strains using synthesis-reasonable satisfaction and multivalue theory merging rule. The genetic diversity of Junzao strains were analyzed by AFLP. 【Result】 Different strains of Junzao showed significant differences in JWB resistance. According to the situations of disease symptom development and phytoplasma infection, the strains were divided into four groups: sensitive, delayed sensitive, delayed resistant and resistant, and 4 strains of Junzao (T15, J2, J5, J8) with very high resistance to JWB were screened out. In the resistant group, the fruit quality of T15, J5 and J8 was also at higher level among the sixteen strains. AFLP analysis showed that there had significant differences in DNA level among different strains of Junzao. 【Conclusion】 Four strains of Junzao (T15, J2, J5, J8) with very high resistance to JWB were screened out, and three of them (T15, J5 and J8) also have an excellent fruit quality.

A Construction of the Core-Collection of Juglans regia L. Based on AFLP Molecular Markers

WANG Hong-Xia-1, ZHAO Shu-Gang-2, GAO Yi-3, XUAN Li-Chun-4, ZHANG Zhi-Hua-1

Scientia Agricultura Sinica. 2013, 46(23): 4985-4995. doi:10.3864/j.issn.0578-1752.2013.23.015

Abstract ( 375 )

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【Objective】 Walnut is an important economic tree species in China, and its germplasm resources are very rich. There were many problems in the processing of germplasm preservation such as large area occupied, higher management costs, which brought great difficulties to the conservation, utilization and evaluation. Therefore, construction of walnut core collection is very important for preservation, study and application of Juglans regia L. germplasm resources.【Method】 Based on AFLP markers, candidate core collections were constructed by using proportional strategy and UPGMA clustering sampling method within subgroups after dividing 131 Juglans regia germplasm into 45 subgroups. The core collection was eventually identified by comparison of the genetic diversity parameters of these candidate core collections, such as the number of polymorphic loci, percentage of polymorphic loci, and morphological indicators. 【Result】The core collections including Tianqiao1 from Hebei province, Xiluo2 and Xilin1 from Shaanxi province, Jinglong1 from Shanxi province, Fenghui from Shandong province, Liaoning8 and Liao73013 from Liaoning province, Wen185 from Xinjiang, Lvbo from Henan province, Beijing746 from Beijing, Vina from America, Qingxiang from Japan, Anbian1 from North Korea, reserved 10% samples of original collection. The retention rate of polymorphic loci was 75.4% in the core collection.【Conclusion】The core collection which was constructed in this study can largely represent the genetic information of the original germplasm, and it was matched with the requirements of the core collection.

A Preliminary Study on Intergeneric Cross-Incompatibility Between Pistacia vera L. and Pistacia chinensis Bunge

LI Xu-Xin-12, CHANG Yue-Xia-1, QU Ping-3, GUO Zhi-Tao-4, CHEN Fang-4, BAI Zhi-Ying-3, LU Bing-She-45

Scientia Agricultura Sinica. 2013, 46(23): 4996-5002. doi:10.3864/j.issn.0578-1752.2013.23.016

Abstract ( 402 )

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【Objective】 The objective of this experiment is to investigate the fertilization barriers of the intergeneric cross between P. vera and P. chinensis. The pollen morphology, the pollen viability and the progress of cross-pollination and fertilization were observed and the intergeneric cross compatibility between P. vera and P. chinensis was studied. 【Method】 The pollen morphology of male parents was observed by scanning electron microscopy. The pollen germination and the viability were tested by means of in vitro cultivation and the progress of pollen tube growth was observed by fluorescence microscopy. 【Result】There are two and four apertures in pollen surface of P. vera and P. chinensis, respectively, and more dense thorn-like particle in P. chinensis pollen grain than in P. vera. The length of polar axis and equatorial axis of P. chinensis pollen grain were 53.84 μm and 44.85 μm, respectively, which is slightly longer than P. vera pollen grain. The pollen germination rate of P. chinensis Bunge and T-491 were 77.30% and 84.38% respectively, and the pollen viability of P. chinensis Bunge and T-491 were 82.4% and 89.14%, respectively. The pollen of P. chinensis begin to germinate at 4 h, the pollen tubes reached ovary at 48 h and enter embryo sac at 72 h in interspecific-cross after pollination, while the pollen of P. vera T-491 begin to germinate at 2 h, the pollen tubes reached the base of stigma at 12 h and reach to ovary at 24 h. The incompatibility characters such as the pollen tubes bending, swollen and lots of irregular callose depositing in stigma and style were observed in interspecific cross. The percentage of final fruit set was 57% in the self-pollination, while the value was 26% in interspecific-cross which is lower than the former.【Conclusion】A few pollen tubes of P. chinensis could enter the style of P. vera and reached to the ovary. There were some pollen tube elongation growth barriers at interspecific-cross between P. vera and P. chinensis. Abnormal pollen tube growth after pollination may be one of the reasons led to low fruiting rate in intergeneric cross between P. vera and P. chinensis.

Analysis of the Genetic Relationship of the Main Cultivars of Ping’ou Hybrid Hazelnut (C. heterophylla×C. avellana) by FISH-AFLP Markers

MA Qing-Hua-1, CHEN Xin-2, ZHAO Tian-Tian-1, LIU Qing-Zhong-2, WANG Gui-Xi-1

Scientia Agricultura Sinica. 2013, 46(23): 5003-5011. doi:10.3864/j.issn.0578-1752.2013.23.017

Abstract ( 338 )

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【Objective】The objective of this study is to establish FISH-AFLP analysis system for Ping’ou hybrid hazelnut and analyze the genetic relationship of the main cultivars.【Method】Ten main cultivars, such as Dawei, were selected as materials. High-quality genomic DNA was extracted with improved CTAB method, and the purified DNA samples were digested with PstⅠ/MseⅠ. After ligation reaction, the samples were used to perform FISH-AFLP detection. The information of all bands was converted into “0-1” matrix. NTSYS pc 2.11F and Popgene 1.32 software were used to conduct the data analysis and plotting.【Result】The results showed that: 15 PstⅠ/MseⅠ primer pairs were selected from 64 combinations with high polymorphism, strong signals and clear bands. A total of 1 739 bands were obtained. The average percentage of polymorphic bands was 97.94%. The similarity coefficient of the main cultivars varied from 0.7556 to 0.8543. All the samples were divided into 4 groups when the threshold value was 0.8398. Each in cutivars of Yuzhui (84-310), Liaozhen No. 4 (85-41) and Ping’ou 69 (84-69) formed an independent group alone and the others constituted the fourth group. The effective number of alleles, Nei’s gene diversity and Shannon’s information index of the main cultivars were 1.3921, 0.2482 and 0.3957, respectively, indicating high genetic diversity among Ping’ou hybrid hazelnut. The unique bands obtained in the experiments could be used in the rapid identification of the main cultivars of Ping’ou hybrid hazelnut. 【Conclusion】 Due to the complex selecting group, Ping’ou hybrid hazelnut is a high polymorphic population and the genetic relationships of the main cultivars are complex. The FISH-AFLP analysis system, PstⅠ/MseⅠprimer pairs and all the genetic parameters obtained in this research can offer reference for the relative researches in Corylus.

Determination of Clavulanic Acid Residue in Edible Tissues of Animals by High Performance Liquid Chromatography-Tandem Mass Spectrometry

LI Shuai-Peng, HUANG Xian-Hui, YANG Gang, GUO Chun-Na

Scientia Agricultura Sinica. 2013, 46(23): 5012-5019. doi:10.3864/j.issn.0578-1752.2013.23.018

Abstract ( 459 )

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【Objective】A confirmatory method was developed for the determination of clavulanic acid (CLAV) residue in edible tissues of animals by high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). 【Method】 One gram tissues were extracted by 3 mL 0.01 mol•L-1 ammonium acetate solution and 3 mL acetonitrile，a back-extraction of the acetonitrile with dichloromethane，aqueous layer was diluted to 4mL，and the sample was determined by HPLC-MS/MS after defatted with N-hexane. The chromatographic separation was achieved on a Luna 5μ C8 column using 0.1% formic acid in water and acetonitrile as mobile phases with gradient elution. The identification of CLAV was carried out by MS/MS equipped with electrospray ionization in negative scanning and multiple reaction monitoring (MRM) modes. 【Result】Matrix-matched calibration standard was used for the quantification. Good linearity was achieved with correlation coefficients more than 0.999 for all analytes in the concentration of 5-500 μg•L-1. The limit of detection (LOD，S/N≥3) was 10 μg•kg-1 in tissues, and the limit of quantification (LOQ，S/N≥10) was 20 μg•kg-1. The mean recoveries at the four spiked levels of LOQ, 1/2MRL (the maximum residue limit), MRL and 2MRL were in the range of 76.39%-89.26% , with the inter-day relative standard deviations (RSD) of 1.89%-6.20%. 【Conclusion】In conclusion, the established method can be applied for the determination of CLAV residues in edible tissues of animals.

Genetic Parameter Estimation for Body Measurements and Weight at Birth in Sanhe Cattle

REN Xiao-Li-1, ZHANG Xu-1, WANG Ya-Chun-1, WU Hong-Jun-2, LIU Ai-Rong-3, ZHANG Yi-1, WANG Dong-Sheng-2, CUI Jiu-Hui-3, DOU Tong-Xi-2, YUAN Peng-2, JIANG Li-Xin-2, ZHOU Lei-2, ZHAO Jian-2

Scientia Agricultura Sinica. 2013, 46(23): 5020-5025. doi:10.3864/j.issn.0578-1752.2013.23.019

Abstract ( 401 )

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【Objective】 In this study, body measurements at birth of 5521 Sanhe cattle born from 2000 to 2012 were collected for genetic parameters estimation for height at withers, body length, chest girth and cannon circumference at birth and birth weight.【Method】DMU software was used, and AI-REML combined EM algorithm based on multiple-trait animal model was employed including sex and age of dam as fixed effects, birth herd-year-month and additive animal genetic effect as random effects.【Result】The results showed that the heritability estimates for height at withers, body length, chest girth, cannon circumference and birth weight were 0.46, 0.48, 0.38, 0.50 and 0.41, respectively. Genetic correlations among these traits varied from –0.05 (between cannon circumference and body length and birth weight, respectively) to 0.86 (body length and birth weight). Phenotypic correlations between these traits ranged from -0.10 (body length and cannon circumference) to 0.70 (body length and birth weight). 【Conclusion】 The heritability estimates were between 0.38 and 0.50 for body measurements at birth and birth weight of Sanhe cattle, which are high inheritable. These traits can be directly selected according to their breeding values, or use indirect selection on traits that are difficult to measure but genetically correlated with other traits (e.g. genetic correlation between 0.40-0.86), for further improvement of breeding efficiency and accuracy.

Selection of Effective SREBP1 shRNA in Cattle and the Construction of Recombinant Adenovirus Vector

FU Chang-Zhen-1, ZAN Lin-Sen-12, WANG Hong-1, CHENG Gong-12, WANG Hong-Bao-12, LI Yao-Kun-1, JIANG Bi-Jie-1, GAO Jian-Bin-1, YANG Ning-1

Scientia Agricultura Sinica. 2013, 46(23): 5026-5036. doi:10.3864/j.issn.0578-1752.2013.23.020

Abstract ( 358 )

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【Objective】 The aim of this study was to construct recombinant adenovirus carrying effective small hairpin RNA (shRNA) which can exclusively interfere NotI bovine sterol-regulatory-element-binding protein 1 (SREBP1) gene expression, thus providing a basis for studying the function and mechanism of SREBP1 gene at cellular level.【Method】According to the coding sequence (CDS) region of SREBP1 gene, six pairs of inhibition shRNA and one pairs of negative control shRNA were designed and further inserted into pENTR-U6 to construct pENTR-U6-shRNA expression vector. The cotransfection of expression vector psiCHECK-Ⅱcarrying SREBP1 and the obtained pENTR-U6-shRNA was carried out in 293 A cell lines to select efficient shRNA. The efficient shRNA and shRNA-NC were connected to pAD/BL-DEST to construct the recombinant plasmid, respectively. The obtained recombinant adenovirus vectors were transfected into 293A cells to package. Then, the adenovirus were amplified and harvested. Real-time PCR (qRT-PCR) was used to detect and confirm the interference effect of the harvested adenovirus on the target gene SREBP1 in bovine pre-adipocytes. The viral titer was determined by GFP labeling method. 【Result】Results showed that shRNA-1053 significantly decreased the expression of SREBP1 by 87.4%. The linearized recombinant adenovirus vector carrying shRNA-1053 and shRNA-NC transfected 293A cells, and further packaged and amplified high-titer recombinant adenovirus Ad-1053 and Ad-NC (7×108 GFU/mL and 9×108 GFU/mL). qRT-PCR results elucidated Ad-1053 significantly down-regulated mRNA expression level of SREBP1 gene in bovine pre-adipocytes (Interfering efficiency ＞85%), while Ad-NC did not.【Conclusion】In this study, the recombinant adenovirus, carrying efficient shRNA designed for RNA interference study of bovine SREBP1 gene, was constructed successfully.

Ovarian Immunohistochemistry Localization and Preantral Follicles Expression Pattern Analysis of Buffalo Proliferating Cell Nuclear Antigen

MA Fan, ZHOU Yu, GAO Ya-Ke, ZHU Peng, LEI Xiao-Can, LIU Qing-You, SHI De-Shun

Scientia Agricultura Sinica. 2013, 46(23): 5037-5043. doi:10.3864/j.issn.0578-1752.2013.23.021

Abstract ( 409 )

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【Objective】To explore the involvement of nuclear antigen protein (PCNA) duiring different periods of buffalo follicular genesis, the expression pattern and ovary immunohistochemistry localization was performed in the present study.【Method】The localization of PCNA was demonstrated in ovarian tissue through the method of paraffin sections with immunohistochemistry, the mRNA expression level of PCNA in preantral follicles of buffalo was determined by real-time fluorescence quantification PCR(QRT-PCR).【Result】The immunohistochemical results showed that, in primordial follicle, no remarkable staining for PCNA in granulosa cells was observed, except the staining of nuclei in oocyte. In primary to secondary follicle, positive staining in oocytes and in some granulosa cells was observed, and PCNA immunoreactivity in granulosa cells of secondary follicle was obviously enhanced than that of primary follicles.The antral follicles showed extensive PCNA labeling in the layers of granulosa and theca cells and in the cumulus cells encircling the oocyte. The results of QRT-PCR showed that,the expression of PCNA in primordial follicles, primary follicles and secondary follicles showed a upregulation trend (43.50333±0.138338 vs 1.633804±0.093796 vs 1±0.012219 , P＜0.01).【Conclusion】In buffalo primary follicles, the expression of PCNA could be detected in granulosa cells, and was obviously enhanced following the development of follicle. The expression pattern of PCNA determined by QRT-PCR in preantral follicles was accordant with the results of immunohistochemistry. These results will provide a foundation for clarifing the mechanism of PCNA involving of follicular genesis in Buffalo.

Distribution of Luteinizing Hormone/Choriogonadotropin Receptor in the Stellate Ganglia of Goat and Its Implications

FAN Jie, CHEN Wen-Dong, DONG Wei, XU Yong-Ping, GUO Xiao, JIN Xiu-Fang

Scientia Agricultura Sinica. 2013, 46(23): 5044-5049. doi:10.3864/j.issn.0578-1752.2013.23.022

Abstract ( 372 )

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【Objective】 The present study was carried out to examine the existence of LCGR (luteinizing hormone/ choriogonadotropin receptor, LCGR) in the stellate ganglion (SG) of goat. 【Method】 Five pairs of stellate ganglions (SG) were taken from mature female and male goats. The distribution characteristics of LCGR were studied after immunohistochemical SP staining. Image-Pro Plus 6.0 (IPP 6.0) was used as the method of semi-quantitative image analysis to evaluate the expression difference of LCGR between neurons and non-neuron cells. 【Result】Neurons and non-neuron cells in SG were immunolabelled in different degrees. In the neurons, uniformly strong immunostaining was found in the whole cytoplasm and neurites, whereas no receptors were detected in the nucleus. In the non-neuron cells including satellite cells and vascular endothelial cells, the cytoplasm was weakly immunolabelled. A significant difference in the intensity of immunolabelling could be found between neurons and non-neuron cells. 【Conclusion】 LCGR was widely distributed in SG of goat and mainly in the cytoplasm and on the membrane of neurons, suggesting that LH and CG may exert potential effects on SG.

Phosphoproteome Analysis of Hypopharyngeal Glands of High Royal Jelly Producing Bee (Apis mellifera L.)

LU Xiao-Shan-12, HAN Bin-2, ZHANG Lan-2, FENG Mao-2, FANG Yu-2, LI Rong-Li-2, ZHOU Tian-E-12, LI Jian-Ke-2

Scientia Agricultura Sinica. 2013, 46(23): 5050-5057. doi:10.3864/j.issn.0578-1752.2013.23.023

Abstract ( 390 )

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【Objective】High royal jelly producing bee (Apis mellifera L.) is the unique honeybee resource in China. However, the mechanism of high royal jelly producing has not been clearly addressed. In order to reveal the significance of protein phosphorylation in hypopharyngeal gland for royal jelly synthesis and secretion, the phosphoproteome of hypopharyngeal gland of nurse bee (6-12 day) was analyzed.【Method】IMAC (immobilized metal-affinity chromatography) phosphoprotein enrichment, SCX (strong cation exchange) separation, LC-MS/MS (liquid chromatography-mass/mass) identification and bioinformatics analysis were applied to analyze phosphoproteome of hypopharyngeal gland of nurse bee.【Result】Of the identified 117 proteins in the hypopharyngeal gland, 6 of them were phosphorylated on 6 phosphopeptides and assigned 8 phosphorylated sites. They were related to protein translation and synthesis, such as 60S acidic ribosomal proteins P0, P1, P2 and 60S ribosomal protein L15, and major royal jelly proteins 1 and 7 precursor.【Conclusion】The phosphorylation modifications occurred on ribosome proteins of hypopharyngeal gland mainly contribute to the high efficiency of synthesizing and secreting of royal jelly protein. The phosphorylation of royal jelly proteins 1 and 7 maintain the reasonable ratio of calcium to phosphorus of royal jelly with the increasing yield, thus meeting the nutrition demand of fertile egg-laying queens and developing larvae. Hence, the data obtained in this study will provide new knowledge to deeply understand the mechanism how high royal jelly producing bees could produce higher amount of royal jelly at the level of protein phosphorylation.

cDNA-AFLP Analysis of Differentially Expressed Resistant Genes of Minn2761

LUAN Zhao-Jie, CAO Yuan-Yin, LI Tian-Ya, CHEN Si, CHEN Xiu-Mei, ZHU Gui-Qing, LI Wei-Hua

Scientia Agricultura Sinica. 2013, 46(23): 5058-5065. doi:10.3864/j.issn.0578-1752.2013.23.024

Abstract ( 374 )

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【Objective】The objective of this study is to analyze the differential genes expression of the important Chinese supplementary differential Minn2761 induced by wheat stem rust, and to find out the fragments related to disease-resistant gene expression. 【Method】 The study was carried out by cDNA-AFLP analysis of differentially expressed resistant genes in Minn2761 and Thatcher inoculated with wheat stem rust.【Result】About 7 000 bands were amplified by 140 primer combinations and by average 50 bands appeared per pair of primers. Through cDNA-AFLP analysis, 86 primer combinations amplified specific bands between Minn2761 and Thatcher. Thirty-five possibly disease resistance-related transcript derived fragments (TDFs) were cloned and sequenced. BLASTx analysis showed that the sequences of 30 TDFs were found to be homologous with those in GenBank. Among them, 24 TDFs had known function and 6 TDFs had unknown function.【Conclusion】Eight TDFs having a highly homology with known genes which encoded kinase binding protein, NBS-LRR resistance protein, cinnamoyl CoA reductase-like 2b, cinnamoyl CoA reductase-like 2c, sucrose-phosphate synthase 7, metacaspase-1-like isoform 1, Zn-finger-like protein, and putative disease resistance protein RGA4-like were supposed to the resistance to stem rust of Minn2761.

Energy Requirements of Fattening Dorset and Thin-Tailed Han Crossbred Ram Lambs

PENG Jin-Jin-12, ZHANG Ying-Jie-1, LIU Yue-Qin-1, JIA Shao-Min-1, DU Wei-Jia-1

Scientia Agricultura Sinica. 2013, 46(23): 5066-5074. doi:10.3864/j.issn.0578-1752.2013.23.025

Abstract ( 417 )

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【Objective】This research aimed to define the maintenance and growth of net energy and metabolic energy requirements parameters for Dorset sheep and thin-tailed Han sheep crossbred ram lambs in the fattening period, to provide a theoretical basis for scientific feeding mutton lambs.【Method】Thirty-five Poll Dorset sheep and thin-tailed Han sheep crossbred non-castrated ram lambs at 6 months of average age and (34.54±0.40) kg of initial body weight were selected in the study, and twenty-five lambs of them were used in a comparative slaughter trial. They were randomly divided into 3 intake level groups which were ad libitum group(15n),60% of ad libitum group(5n) and 40% of ad libitum group(5n), and ad libitum group was divided into three groups, each group have five sheep. Forth similar weight lambs of ad libitum 1st group, ad libitum 2nd group and ad libitum 3rd group were slaughtered on 1st day, 19th day(43 kg of average body weight) and 63rd day (50 kg of average body weight), respectively. Meanwhile, forth similar weight lambs of 60% and 40% of the ad libitum group were slaughtered concomitantly with lambs of ad libitum 3rd group. Another nine lambs were used in the digestion-metabolism trial, the lambs were randomly divided into three intake level groups as comparative slaughter trial. Total collection of feces and urine was conducted, and the recommended formulation was used to predict the methane energy. 【Result】The calculated daily net energy for maintenance (NEm) was 286.88 kJ•kg-1 metabolic empty BW (EBW0.75) or 269.09 kJ•kg-1 metabolic BW(BW0.75). The daily metabolizable energy for maintenance (MEm) was 423.89 kJ•kg-1 metabolic empty BW (EBW0.75) or 397.60 kJ•kg-1 metabolic BW(BW0.75). Net energy and metabolizable energy requirements for growth respectively ranged from 1.45 to 6.20 MJ•d-1 and 3.21 to 13.72 MJ•d-1 for daily gains of 100-350 g BW. The partial efficiency of ME utilization for maintenance (km) were 0.68. The partial efficiency of ME utilization for growth (kg) were 0.45. 【Conclusion】Energy requirement parameters of Dorest sheep and thin-tailed Hansheep ram lambs in the fattening period had some differences with those reported by NRC(2007).

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