Scientia Agricultura Sinica

Genetic Diversity and Genome-Wide Association Analysis of Stripe Rust Resistance Among the Core Wheat Parent Zhou 8425B and Its Derivatives

XIAO Yong-Gui, YIN Gui-Hong, LI Hui-Hui, XIA Xian-Chun, YAN Jun, ZHENG Tian-Cun, JI Wan-Quan, HE Zhong-Hu

Scientia Agricultura Sinica. 2011, 44(19): 3919-3929. doi:10.3864/j.issn.0578-1752.2011.19.001

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【Objective】 Zhou 8425B is the most important core parent and has been widely used in the South Part of the Yellow and Huai Valleys Winter Wheat Zone. Understanding the principal components and detecting the wheat stripe rust resistance loci in Zhou 8425B will provide important information for the future genetic improvement of wheat varieties. 【Method】 The core parent Zhou 8425B and its 50 derivative varieties and lines were used to compare their genetic structure and components among different generations. Whole-genome association mapping was employed to identify the stripe rust resistance loci using 921 Diversity Array Technology (DArT) and 83 SSR markers. 【Result】The averaged genetic similarity index was 67.6% among Zhou 8425B and its derivatives, and the clustering results for these genotypes were basically in consistent with that from the pedigree analysis. On the whole-genome levels, Zhou 8425B had the contribution ratios of 67.7%, 63.6% and 58.8% to its first, second and third generation derivatives, respectively. The genetic contribution ratios of A, B and D subgenome levels were 68.7%, 62.0% and 59.4%, respectively. As for single chromosome level, 44.9% (4A) to 70.9% (1D) of Zhou 8425B alleles could be detected on the derivative varieties. Four loci were detected to be associated with stripe rust resistance by association mapping (P＜0.01), in which two loci QYr.caas-2BL and QYr.caas-7BL were found in the same genomic regions of Yr7 and YrZH84, respectively. A QTL QYr.caas-1BL was found in the same genomic region as leaf rust resistance gene LrZH84 reported previously, which might be attributed to be pleiotropic effect or tight linkage each other. A new locus QYr.caas-3AL was located on the long arm of chromosome 3A and linked with the terminal marker wPt-0398, explaining 22.9% of the phenotypic variance.【Conclusion】The contribution ratios transferred from the core parent to its derivatives on genome and chromosome levels were probably caused by various important alleles associated with agronomic traits and resistance genes. Four QTLs for stripe rust resistance detected in 50 derivative varieties were derived from the core parent Zhou 8425B. Therefore, it is presumed that these loci for stripe rust resistance and other important traits derived from Zhou 8425B could play important roles in the future wheat breeding program in China, especially in the South Part of the Yellow and Huai Valleys Winter Wheat Zone.

Cloning and Activity Analysis of Protein KinaseTaNPK Gene Promoter in Wheat

DI Chao-Zeng, XU Zhao-Shi, CHEN Yao-Feng, LIU Pei, LI Lian-Cheng, CHEN Ming, MA You-Zhi

Scientia Agricultura Sinica. 2011, 44(19): 3930-3936. doi:10.3864/j.issn.0578-1752.2011.19.002

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【Objective】 Cloning the promoter of a plant protein kinase gene, TaNPK, and analyzing its response to salt stress, could be helpful to investigate the regulatory mechanism of TaNPK gene and to provide a theoretical foundation for analyzing salt-tolerance mechanism of wheat. 【Method】 Genome walking technology was used to amplify the upstream regulatory sequence of TaNPK gene, and six different 5′UTR deletion mutants of the TaNPK gene promoter were amplified by PCR, and then inserted into the vector pBI121 to replace CaMV 35S promoter respectively. The recombinant plasmids were transferred into Arabidopsis leaf protoplasts by PEG-mediated transient expression system and the promoter activities were quantitatively estimated using gus report gene. 【Result】 A 2 004 bp 5′flanking sequence was obtained by genome walking technology. Deletion analysis was made by comparing with the control. The results coincided with that the GUS gene driven by six 5′-end deletion mutants could be highly effectively expressed in protoplasts, and GUS activity increased in varying degrees with the treatment of NaCl compared with real-time fluorescent quantitative PCR analysis. 【Conclusion】 The TaNPK gene promoter was cloned, the activity analysis showed that NaCl up-regulates TaNPK gene in wheat, and a negative regulatory factor maybe exist between -1 083—-1 296 bp area of the 5′UTR of TaNPK gene.

The Relationship on Anther Tapetum, Sporopollenin and Expression of RAFTIN1 in Physiological Male Sterile Wheat

SHENG Ying, ZHANG Gai-Sheng, LI Ya-Xin, ZHANG Long-Yu, WANG Shu-Ping, ZHAO Xin-Liang, WANG Liang-Ming, SONG Yu-Long

Scientia Agricultura Sinica. 2011, 44(19): 3937-3944. doi:10.3864/j.issn.0578-1752.2011.19.003

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【Objective】 To pave the way for revealing the mechanism of physiological male sterility in wheat, the relevance among expression of RAFTIN1, anther tapetum and sporopollenin were studied.【Method】Using physiologically male sterility induced with chemical hybridizing agent SQ1, genetically male sterility and corresponding normal lines of complete fertility as experimental materials in wheat, morphological changes of anther tapetum from different developmental stages of microspore, sporopollenin accumulation and RAFTIN1 expression were analyzed via paraffin section, cell fluorescence staining and Real-time PCR. 【Result】The results showed that anther tapetum of physiological male sterile lines pre-degraded, sporopollenin secretion decreased, while RAFTIN1 expression level was maximum at mononuclear stage, and contributed to a large number of sporopollenin to pollen intine. At binuclear and trinuclear stages, the tapeta of physiological male sterility were degraded completely, sporopollenin secretion was ceased and expression patterns of RAFTIN1 were down-regulated significantly. 【Conclusion】 These data demonstrated that the mechanisms of physiologically male sterility induced by SQ1 were associated with pre-degradation of tapetum and expression degree of RAFTIN1 with responsibility for transporting sporopollenin.

Genome-Wide Identification, Classification and Expression Analysis of LEA Gene Family in Soybean

LI Le, XU Hong-Liang, YANG Xing-Lu, LI Ya-Xuan, HU Ying-Kao

Scientia Agricultura Sinica. 2011, 44(19): 3945-3954. doi:10.3864/j.issn.0578-1752.2011.19.004

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【Objective】 Identification, phylogenetic and tissue specific expression pattern analysis of LEA family genes in Soybean will be helpful to study the functions of plant LEA genes. 【Method】 Based on soybean genome database and bioinformatic method, soybean LEA family genes were identified and their gene sequences, chromosome location and duplication information were obtained. LEA proteins were classified according to their structure and phylogeny relationship. The soybean developmental expression database and EST data from NCBI UniGene were used to analyze the expression pattern of these genes at different development stages.【Result】A total of 36 LEA genes were systematically identified from soybean and classified into 8 subfamilies according to motif analysis and phylogeny relationship. They were located on 16 chromosomes and most of them had stress related cis-acting elements in their promoter regions. Ten differentially expressed genes were found, and five of them were highly expressed in seeds by microarray and EST data at each developmental stages.【Conclusion】Thirty six LEA family genes in soybean were identified by genome-wide screening. They were classified into 8 subfamilies and distributed on 16 chromosomes with stress related cis-acting elements in their promoter regions. They were differentially expressed in different tissues and developmental stages. These results are helpful for their functional analysis and utilization in crop genetic improvement.

作物精确栽培；栽培方案设计；生长指标诊断；生产力预测Development and Implementation of Crop Precision Cultivation Technology

CAO Wei-Xing, ZHU Yan, TIAN Yong-Chao, YAO Xia, TANG Liang, LIU Xiao-Jun, NI Jun

Scientia Agricultura Sinica. 2011, 44(19): 3955-3969. doi:10.3864/j.issn.0578-1752.2011.19.005

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The objective of this paper is to introduce the technical content, research progress, application impact, and future prospect of crop precision cultivation (CPC). The contents of CPC include digital design, information sensing and dynamic simulation of the objects and processes of crop cultivation system and thus realizing quantitative and precise crop management, based on integration of system science, information technology and crop cultivation. During the recent years, Chinese-style CPC technology has been developed by thorough and systematic research on the key techniques and application systems of crop precision cultivation, and remarkable advances have been achieved in three areas of CPC technology as precision design of crop cultivation plan, precision diagnosis of crop growth status, and precision prediction of crop productivity. Among them crop cultivation plan design is based on local production factors, crop growth monitoring and diagnosis is based on canopy spectral reflectance, and crop yield and quality prediction is based on growth process models. The future prospect of CPC is directed toward integrating and coupling crop cultivation with simulation technique, sensing technique, and decision support technique, and improving the key techniques for design of crop cultivation plan, diagnosis of crop growth status, and prediction of crop productivity from the field to regional scales. Further, the comprehensive software and hardware systems on the basis of CPC technology should be constructed and applied for realizing informatization of crop production management and promoting development of smart agriculture. Crop precision cultivation will be helpful to enhancement of the management level and production output, and facilitation of the progress of informatization and modernization of agricultural industry.

A Difference in Source-Sink Structure and Chalky Grain in Early Indica Rice Cultivars with Different Sink-Potentials

XIONG Jie, GENG Chun-Miao, DING Yan-Feng, WANG Shao-Hua

Scientia Agricultura Sinica. 2011, 44(19): 3970-3980. doi:10.3864/j.issn.0578-1752.2011.19.006

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【Objective】 The aim of this study is to reveal the effect of source-sink structure at full heading stage to chalky grain of early indica rice cultivars with different sink potentials. 【Method】 Eighteen hybrid early rice cultivars were cultured in 2008 and 2009, respectively. The tested rice cultivars were classified into 3 types with large, middle and small sink potential respectively by using cluster analysis method. Then source and sink structure, grain filling characteristics and chalky grain of each sink potential types were compared, based on which the effects of source-sink structure on grain filling and chalky grain were analyzed. 【Result】 The results showed that LAI and net assimilation rate during grain filling were in the order of large sink potential type＞middle sink potential type＞small sink potential type. But comparisons of leaf area, stem and sheath weight and net assimilation rate per unit sink potential showed an opposite order. Grain filling rate was in the order of large sink potential type＜middle type＜small type, while grain filling duration showed an opposite order. Chalky grain rate and chalkiness were in the order of large type＞small type＞middle type. 【Conclusion】 As large and middle sink potential type cultivars had inadequate grain filling material, low grain filling rate, high chalky grain rate and chalkiness, strategies for enlarging source size should be used to lower chalky grain; And as small sink potential type cultivars had sufficient grain filling material, high grain filling rate, high chalky grain rate and chalkiness, strategies for enlarging sink size should be used to lower chalky grain.

Effect of Aromatic Plant-Derived Nutrient Solution on Inhibition of Harmful Bacteria and Nutrition for Pear Plants

LIU Yan-Na, CUI Nan-Nan, ZHANG Jie, YU Feng-Ming, ZHANG Li-Bin, YAO Yun-Cong

Scientia Agricultura Sinica. 2011, 44(19): 3981-3990. doi:10.3864/j.issn.0578-1752.2011.19.007

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【Objective】 The objective of this study is to provide a simple, low cost, and effective plant nutrition agent for organic fruit production and explore the bacteriostat effects of aromatic plant-derived nutrient solution on Venturia pirina Aderh, Pnysalospora pinicola Nose and Valisa mali vegetative growth and blade nutrient content. 【Method】 Biennial potted nanshui pear Pyrus pyrifolia (Burm. f.) Nakai/Pyrus betulaefolia Bunge trees were selected as materials for treatments of foliar application with plant-derived nutrient solutions. The treatments were Tagetes patula (Tr1), Perilla frutescens (Tr2), Hyssopus officinalis (Tr3), Mentha canadensis (Tr4), Centaurea cyanu (Tr5), as well as their proportional mixture (Tr6), respectively. And bio-bacterial manure Nanguochun, a kind of foliar fertilizers (Tr7), and water (Tr8) were used as control. 【Result】 The results indicated that the foliar application of aromatic plant-derived nutrient solution and indoor antimicrobial tests was good in bacteriostatic effect of Venturia pirina Aderh, Pnysalospora pinicola Nose and Valisa mali and their anti-bacteria effect respectively reached 76.00%, 82.12%, 87.88% compared with controls. The foliar application of both plant-derived nutrient solutions and Nanguochun promoted the growth of shoots, increased the leaf area and improved the contents of mineral elements in leaves except for element P. In terms of comparison of the effects among different treatments, spraying Tr3 largely improved the growth of pear plants Pyrus pyrifolia (Burm. f.) Nakai/ Pyrus betulaefolia Bunge. 【Conclusion】 It is suggested that foliar application of plant-derived nutrient solution could result in improving the nutrient contents in leaves of trees and the status of nutrition growth, and simultaneously play a role in preventions of Venturia pirina Aderh, Pnysalospora pinicola Nose and Valisa mali in leaves of pear plants.

Direct PCR Detection of Radopholus similis from Plant Tissues and Mixed Sample of Nematodes by ITS Specific Primers

LIU Yi-Fan, XU Chun-Ling, ZHANG Chao, SU Xiu-Min, XIE Hui

Scientia Agricultura Sinica. 2011, 44(19): 3991-3998. doi:10.3864/j.issn.0578-1752.2011.19.008

Abstract ( 863 )

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【Objective】 The objective of this study is to build an ITS-PCR (internal transcribed spacer-polymerase chain reaction) method for detecting Radopholus similis (Cobb) Thorne from mixtures of different nematodes, or tissues of Musa spp. and Anthurium andraeanum. 【Method】 A specific primer pair was designed according to ITS region of rDNA from R. similis using Primer Premier 5.0. A single-step PCR method was used to specially detect the target ITS-rDNA of R. similis. 【Result】 The specificity of the designed primers was confirmed that a 518 bp PCR product was amplified from 7 populations of R. similis, not from the other 16 species including 17 populations. Using the established DNA extraction method and PCR system by the specific primers, the target ITS-rDNA fragment was specially amplified from DNA templates of the mixture of individuals of R. similis with species of Pratylenchus, Helicotylenchus, Rotylenchulus, Meloidogyne, Ditylenchus, Tylenchorhynchus, Hoplolaimus, Filenchus, Aphelenchoides, and Rhabditida, and also from the DNA mixtures of at least 3 individuals of R. similis with root tissues (2 cm long, about 0.1 g) of banana and Anthurium andraeanum. 【Conclusion】 The ITS-PCR method established in this study was proven to be a reliable and useful technique for fast and accurate detecting of R. similis from mixed nematode samples and mixtures of root tissues of banana or A.andraeanum.

Assessment of Tillage Effects on Soil Quality for Double-Rice Paddy with Tilth Index

XU Shang-Qi, ZHANG Ming-Yuan, SUN Guo-Feng, TANG Wen-Guang, CHEN Fu, ZHANG Hai-Lin

Scientia Agricultura Sinica. 2011, 44(19): 3999-4006. doi:10.3864/j.issn.0578-1752.2011.19.009

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【Objective】 Improvement of soil quality is very meaningful for increasing land productivity and ecological environment. In this study, tilth index (TI) was used to evaluate the soil quality of double-rice paddy under different tillage systems in order to provide a basis for selecting reasonable tillage. 【Method】 No-tillage (NT), conventional tillage (CT) and rotary tillage (RT) were carried out in 2005 at the Ningxiang County, Hunan province. Soil organic matter (SOM), bulk density (BD), total nitrogen (TN) and pH of 0-20 cm soil were measured to calculate TI, and soil samples were collected at rice harvest time in both 2007 and 2008. TI was calculated by a proposed regression model. 【Result】 Rice yield had a significant negative correlation with bulk density (R2=0.67) and positive correlation with soil organic matter (R2=0.45). TI increased linearly with increasing rice yield (R2=0.67). TI could be easily determined through measured SOM and BD of the soil, TI = 0.40 SOM' + 0.60 BD', the proposed regression model showed the value of TI was a good indicator for optimum rice yields, as well as an evaluation of the soil quality. In this study, soil quality of all the three tillage treatments was obviously improved, the change of TI showed that NT didn’t have a better soil quality at early period of experiment, but the tendency of soil quality improvement was more significant than CT and RT, therefore long-term no-till and straw returning may be a good choice. 【Conclusion】 The value of TI, which was calculated through the proposed regression model, could be a good indicator for rice yields, as well as an evaluation of the soil quality. Soil quality of NT increased the fastest in all the three tillage systems.

Effects of Urease/Nitrification Inhibitor on Soil Nitrogen Leaching Loss in Loamy Fluvo-Aquic Soil

CHUAN Li-Min, AN Zhi-Zhuang, DU Lian-Feng, ZHAO Tong-Ke, LI Shun-Jiang

Scientia Agricultura Sinica. 2011, 44(19): 4007-4014. doi:10.3864/j.issn.0578-1752.2011.19.010

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【Objective】 The objective of this study was to determine the effects of urease/nitrification inhibitors on the transformation and leaching of nitrogen from urea.【Method】A soil column leaching experiment was conducted in a greenhouse to observe nitrogen transformation and leaching loss following the application of urea with a urease inhibitor N-(n-Butyl) thiophosphoric triamide (NBPT), a nitrification inhibitor dicyandiamide (DCD), or both.【Result】When urea-N was applied at 600 kg•hm-2, NBPT and DCD combined or separately application significantly reduced the NO3--N concentration in the leacheate before 24 days, and reached the peak after 30 days. DCD, or DCD and NBPT combined application retarded the peak by 7 days. DCD had a greater effect on reducing nitrogen leaching through the whole monitoring period, the combination of NBPT or DCD and NBPT had a good effect especially in the latter period. For the whole leaching study period, the combination of NBPT, DCD, DCD and NBPT significantly reduced the nitrate accumulation by 11.6%, 13.7% and 17.2%, respectively. 【Conclusion】When fertilizer was applied to a certain extent, nitrate leaching loss can be reduced by applying urea fertilizer with a urease and/or nitrification inhibitor.

Interaction Between Water Retaining Agent and Nitrogen Fertilizers and the Effect of Water and Fertilizer Conservation

GOU Chun-Lin, WANG Xin-Ai, LI Yong-Sheng, DU Jian-Jun, QU Dong

Scientia Agricultura Sinica. 2011, 44(19): 4015-4021. doi:10.3864/j.issn.0578-1752.2011.19.011

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【Objective】 The objective of the experiment is to study the effects of different nitrogen fertilizers on water absorbent characteristics of water retaining agent (WRA), nutrients adsorption by WRA, and the effect of water and fertilizer-conservation when WRA and different nitrogen fertilizers are applied in combination. 【Method】 Water absorbency rates in different solution of urea, ammonium bicarbonate, ammonium sulfate and ammonium chloride were determined by tea bag method. N adsorption capacity was determined by the difference of N amount in the solution before and after adsorption occurred. Maize was used as the tested crop in a pot experiment to study the effect of water and fertilizer conservation when WRA and different nitrogen fertilizers are applied in combination. 【Result】 The results showed that the water absorbent rate of the WRA decreased obviously along with the increase of concentrations of various N fertilizers. The influence of nitrogen fertilizers on water absorbent rate of the WRA was in the order of urea＜ammonium bicarbonate＜ammonium sulfate＜ammonium chloride. While WRA absorbed water, it could absorb the molecules of urea or NH4+ of the fertilizers dissolved in water. The N amounts of absorbed fertilizers increased with the increase of concentrations of fertilizers. However, the rate of N absorbed showed a decrease trend along with the increase of concentrations of fertilizers except urea.WRA and urea or ammonium bicarbonate applied in combination could obviously increase the bioyield, root dry weight, N use efficiency and WUE, as well as net photosynthetic, transpiration rate and stomatal conductance. 【Conclusion】 While WRA absorbed water, it could absorb N nutrient dissolved in water. N fertilizer strongly decreased water absorbent rate of WRA, however, urea was the weakest. WRA and urea applied in combination had the best effect of water and fertilizer control.

Effects of JA Synthesis-Related Genes Spr2 and LePrs on the

ZHANG Li-Ning, CHENG Ji-Hong, YANG Rui, SUN Zhong-Hua, WU Chun-Xia, WANG Shao-Hui

Scientia Agricultura Sinica. 2011, 44(19): 4022-4028. doi:10.3864/j.issn.0578-1752.2011.19.012

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【Objective】 Spr2 and LePrs were related with JA synthesis, the effects of the two genes on the response of tomato to root-knot nematodes were studied. 【Method】 In this paper, JA biosynthetic mutant (spr2 plants) and JA-overexpression transgenic plants (35S::PS plants) of tomato as well as wild tomato species (CM) were used, and the effects of MeJA sprays and grafting on root-knot numbers, PI-II content and Spr2, LePrs transcription levels, before and after inoculating root-knot nematodes, were analyzed. 【Result】 Compared with CM and 35S::PS plants, spr2 plants was more susceptible to root-knot nematodes. Spraying MeJA on tomato leaves could reduce root-knot numbers. Using 35S::PS plants as stock improved grafting tomato resistance to root-knot nematodes. 【Conclusion】 Resistance to nematodes in JA biosynthetic mutant (with mutant Spr2) was reduced, while in JA-overexpression transgenic tomato (with over-expressing LePrs) it was increased. So Jasmonic acid played a role in the response of tomato to root-knot nematodes.

Molecular Cloning of Isopentenyl Transferases Genes Family in Malus domestica Borkh. and a Preliminary Functional Analysis of MdIPT5a

LI Hao, ZHANG Wen, ZHAO Xu-Mian, REN Xue-Qin, ZHU Yuan-Di

Scientia Agricultura Sinica. 2011, 44(19): 4029-4036. doi:10.3864/j.issn.0578-1752.2011.19.013

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【Objective】The aim of this study was to clone a multigene family encoding isopentenyl transferases in Malus domestica, and analyze the biological functions of MdIPT5a, which would provide a basis for in-depth studies of the functions of MdIPTs in cytokinin biosynthesis and genetic transformation. 【Method】The MdIPTs were isolated from apple cultivar ‘Fuji’ by RACE based on the information of apple genome. The subcellular localization of MdIPT5a was transient expressed in onion epidermal cells by the particle bombardment method and in Arabidopsis mesophyll protoplast by the PEG-calcium transfection. MdIPT5a was transferred into tobaccos (Nicotiana tabacum cv. Wisconsin 38) by Agrobacterium-mediated transformation. Transgenic tobaccos were examined by RT-PCR. 【Result】Ten cDNA sequences of MdIPTs were isolated from apple. Among them seven sequences encoding adenylate isopentenyl transferases that functioned in the major pathway of cytokinin biosynthesis shared a conserved domain of GxxGxGK[S, T] motif at N-teminal, and were located in chromosome No. 13, 16, 3, 11, 13, 16 and 6 respectively. They were designated as MdIPT1a, MdIPT1b, MdIPT3a, MdIPT3b, MdIPT5a, MdIPT5b and MdIPT7a. These genes encoded 284 to 370 amino acids without introns. The MdIPT5a-GFP fusion proteins were located in cytoplasm, but not in plastids. Overexpression of MdIPT5a in tobaccos in vitro induced the phenotypes of hardly rooting with more leaves and adventitious shoots. 【Conclusion】 The phenomenon that genes in pairs performed highly homologous occurred in MdIPTs, which was accordant with the theory that 17 chromosomes in the Pyreae were derived from nine ancestral chromosomes. MdIPT5a exhibited the catalytic function in cytokinin biosynthesis.

Characteristics of SSRs Derived from ESTs and Development of EST-SSR Markers in Litchi (Litchi chinensis Sonn.)

SUN Qing-Ming, MA Wen-Chao, MA Shuai-Peng, ZHAO Jun-Sheng, BAI Li-Jun, CHEN Jie-Zhen, CAI Chang-He, XIANG Xu, 欧Liang-Xi

Scientia Agricultura Sinica. 2011, 44(19): 4037-4049. doi:10.3864/j.issn.0578-1752.2011.19.014

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【Objective】 The objective of this study was to characterize litchi SSRs derived from ESTs and to develop EST-SSR markers for possible use of the markers in the studies of litchi genetic diversities, linkage mapping and phylogenetic relationships.【Method】 SSRIT (simple sequence repeat identification tool) was used to search SSRs in 1 331 unigenes obtained from a cDNA library of pericarp cell enlargement period of litchi. EST-SSR primers were subsequently designed using Primer5.0. The validity and revealing power of the primers were tested by PCR amplification of 16 litchi germplasm accessions showing apparent morphological variations and by PAGE analysis of the PCR products. 【Result】 A search of 1 331 Unigene sequences of litchi identified a total of 220 SSRs that occurred in 189 Unigenes. The average SSR density was one SSR per 4.42 kb of EST sequences, and the average SSR length was 18.43 bp. The dinucleotide and trinucleotide repeats, which were 37.27% and 31.82%, respectively, were the most abundant SSRs detected and accounted for 69.09% of the total. Among the 72 observed repeat motifs, GA/CT was the most abundant (16.82%), followed by AG/TC(14.55%), A/T(11.82%)，AT/TA(5.00%) and GAA/CTT (3.64%). The number of repeats in a SSR was quite different with different repeat motifs. Among the 150 EST-SSR primer pairs designed, 122 (81.33%) yielded ideal PCR products, and 100 of them showed polymorphic bands. 【Conclusion】 There were abundant SSR loci in ESTs of litchi, and the efficiency of developing SSR markers from litchi ESTs was high. The study has generated 100 EST-SSR markers which are potentially useful in the analysis of genetic variations of litchi. More SSR markers could be developed from EST-SSRs identified in this study.

The Antifungal Properties of Lactobacillus casei AST18 and Its Application as Adjunct Culture in Yogurt

LI Hong-Juan, LIU Lu, ZHANG Shu-Wen, KONG Fan-Pi, SUN Zhuo, LU Jia-Ping

Scientia Agricultura Sinica. 2011, 44(19): 4050-4057. doi:10.3864/j.issn.0578-1752.2011.19.015

Abstract ( 839 )

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【Objective】The antifungal activities of lactic acid bacteria (LAB) isolated from Chinese traditional fermented products were detected. The selected strain was used as adjunct culture in yogurt to control the fungal pollution.【Method】The production of phenyllactic acid (PLA) was detected with HPLC and the correlation between PLA and the antifungal activities was studied. Then the antifungal properties of Lactobacillus casei AST18 culture supernatants were detected. Lactobacillus casei AST18 was used as adjunct culture in the yogurt production process. Four different inoculums of AST18, 2%, 4%, 6%, 8% and a control group which used the commercial starter only were added in yogurt. After fermentation, sensory evaluation，physical and chemical indicators, the fungal growth conditions were investigated. 【Result】According to the comparison of the production of PLA and the antifungal activities, no relationship was observed between the production of PLA and the inhibition zones. The culture supernatant of Lactobacillus casei AST18 was sensitive towards pH and heat treatment but it was not affected after treatment of trypsin and pepsin. The inoculums of 2% in yogurt could inhibit the growth of Penicillium sp. and extend the shelf life. And it had no significant effect on the flavor and sensory quality.【Conclusion】Lactobacillus casei AST18 can inhibit the growth of filamentous fungi conspicuously. It can be used as an adjunct culture in yogurt to prevent the fungal growth and extend the shelf life.

Continuous Extraction of Saponin and Polysaccharide from Momordica charantia L. and Their Inhibitory Effect on α-glucosidase

WANG Qi, DENG Yuan-Yuan, ZHANG Ming-Wei, ZHANG Rui-Fen, ZHANG Yan, TANG Xiao-Jun, WEI Zhen-Cheng, CHI Jian-Wei

Scientia Agricultura Sinica. 2011, 44(19): 4058-4065. doi:10.3864/j.issn.0578-1752.2011.19.016

Abstract ( 983 )

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【Objective】 The technical process and suitable conditions for continuous extraction of saponin and polysaccharide from Momordica charantia L. by ultrasonic-microwave synergistic extraction were established and to evaluate their inhibition effects on α-glucosidase 【Method】 Quadratic Regression Rotational Combinational Design was applied to optimize the extraction process of saponin and polysaccharide from Momordica charantia L. using continuous ultrasonic-microwave synergistic extraction method. The method with substrate 4-nitrophenol-2-?-D-glucopyranoside(PNPG) was used to measure the α-glucosidase inhibitory effect of saponin and polysaccharide extract. 【Result】 The optimum conditions for saponin and polysaccharide extraction form Momordica charantia L. were as follows: the dried Momordica charantia L. with 13% of water content was grinded to pass a 200 μm sieve; the solvent was 75% enthanol and the ratio of material to solvent was 1:18; ultrasonic-microwave synergistic extraction for 16 min (the power of microwave was 50 W and the ultrasonic frequency was 20 kHz). Under this condition the extraction yield of saponin from Momordica charantia L. was 2.51%. The residue was further dissolved in water of pH 8.0 for extraction of polysaccharide, the ratio of material to solvent was 1:14; the ultrasonic-microwave synergetic extraction time was 23 min (the power of microwave was 50 W and the ultrasonic frequency was 20 kHz) at a temperature of 84℃. Under these conditions, the extraction yield of polysaccharide from Momordica charantia L. was 12.86%. Both of the extracts of saponin and polysaccharide from Momordica charantia L. showed an obvious inhibitory activity against α-glucosidase with values of 1.03 and 10.73 mg•mL-1, respectively.【Conclusion】 The raw materials were utilized effectively and the extraction yield and efficiency were increased using a method of ultrasonic-microwave synergistic extraction. The extracted saponin and polysacchride from Momordica charantia L. showed an obvious inhibition effect on α-glucosidase with promising of blood sugar reducing potentials.

Analysis of Chemical Composition in Amniotic Fluid and Allantonic Fluid of Fetus with Different Body Weights in Huanjiang Mini-Pigs

LIU Jun-Feng, WU Chen, KONG Xiang-Feng, YANG Huan-Sheng, GENG Mei-Mei, YIN Yu-Long, HE Ruo-Gang

Scientia Agricultura Sinica. 2011, 44(19): 4066-4071. doi:10.3864/j.issn.0578-1752.2011.19.017

Abstract ( 982 )

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【Objective】 The present study was conducted to investigate the relationship between body weight (BW) of fetus and biochemical parameters of fetal liquid in Huanjiang mini-pigs in order to illuminate the genesis mechanism, and approach prevention and cure methods of intrauterine growth retardation (IUGR). 【Method】 Seven female mini-pigs with a live BW of 35-40 kg were chosen in this study and raised in seven pens of one pig. At the end of a 45-day feeding period after serviced, the sows were killed by bloodletting of arteria carotis, and the BW of fetus and the weight of placenta and womb were got. The amniotic fluid, allantonic fluid and placenta were collected from two fetus with minimal BW or maximal BW per sow, respectively, for analysis. 【Result】 Results showed that the BW of fetus, placenta weight, womb weight, volume of amniotic fluid plus allantonic fluid in fetus with minimal BW were lower by 23%, 62%, 41% and 39% (P＜0.05), respectively. The concentration of total cholesterol in amniotic fluid was higher by 30% (P＜0.05). The concentrations of serine, alanine and tyrosine in allantonic fluid decreased by 58%, 44% and 29% (P＜0.05), while of lysine, arginine and threonine increased by 25%, 40% and 114% (P＜0.05), respectively. The content of total protein and activity of inducible nitric oxide synthetase (NOS) in placenta were higher by 39% and 70% (P＜0.05), respectively, while of constitutive NOS decreased by 22% (P＜0.05), when compared with the fetus with maximal BW. 【Conclusion】 These findings suggested that there is a close relationship between the BW and biochemical parameters of fetus during early pregnancy in Huanjiang mini-pigs.

Polymorphisms of SLC11A1 Gene and Their Association with Mastitis in Chinese Holstein

GUO Yang, WANG Hong-Mei, HOU Ming-Hai, WANG Chang-Fa, HE Hong-Bin, LU Wen-Fa , ZHONG Ji-Feng

Scientia Agricultura Sinica. 2011, 44(19): 4072-4080. doi:10.3864/j.issn.0578-1752.2011.19.018

Abstract ( 700 )

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【Objective】The polymorphisms of SLC11A1 gene and their correlation to mastitis in Chinese Holstein were investigated for the purpose of providing molecular maker information to facilitate the breeding efficiency of mastitis resistance. 【Method】CRS-PCR, descend PCR and sequencing were applied to analyze the polymorphisms of four locus in SLC11A1 gene intron 9, exon10 and intron 11 in 771 Chinese Holstein Cattle. The PHASE and SHEsis softwares were used to analyze matching chain disequilibrium and haplotype analysis, respectively. 【Result】 Four SNPs not linking were found, in which 6 067(A/G), 6 358(C/T) and 7 809(A/T) were reported for the first time, and were in intron 9, exon 10 and intron 11, respectively. At locus 6 358 (C/T) and 7 155 (A/G), the cows with genotype CC and AA showed higher milk yield and lower SCS (P＜0.05). Sixteen haplotype combinations were found in 771 Chinese Holstein Cattle, the cows with haplotype combinations CAAA had the higher milk yield and lowest SCS.【Conclusion】Considering the distribution frequencies of haplotype combinations, SLC11A1 gene CAAA haplotype combination is favorable haplotype combinations to the SCS and milk yield and can be chosen as the molecular markers of choosing mastitis resistance selection in dairy cattle.

Phylogenetic Relationships Among Domestic Chinese Bovinae Species Based on Mitochondrial Cytochrome b Gene Sequences

GENG Rong-Qing, WANG Lan-Ping, JI De-Jun, CHANG Hong, LI Yong-Hong, CHANG Chun-Fang

Scientia Agricultura Sinica. 2011, 44(19): 4081-4087. doi:10.3864/j.issn.0578-1752.2011.19.019

Abstract ( 658 )

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【Objective】This study was designed to analyze the phylogenetic relationships among domestic Chinese Bovinae species and provide molecular evidence for phylogenetic evolvement.【Method】The complete sequences of mitochondrial cytochrome b gene from six bovine species, including Bos taurus, Bos indicus, Bos grunniens, Bos frontalis, Bubalus bubalis (swamp buffalo) and Bos bubalis (river buffalo), were obtained by PCR and standard double-strand DNA sequencing method. The phylogenetic relationships among different bovine species were analyzed through molecular evolution software.【Result】The complete sequences of mitochondrial cytochrome b gene from six bovine species were all 1 140 bp in length. No insert and deletion mutation was observed. Total 220 variation sites were tested including 213 parsimony informative sites. The number of transition was higher than transversion and the ratio was 5.2, suggesting that the mutation was not saturation. Genetic distances showed that Bos taurus and Bos indicus possessed the most close relationship and the relationships between Bos grunniens, Bos frontalis, swamp water buffalo, river water buffalo and them deduced in turn gradually. Six bovine species distributed four main monophyletic branches and could be divided into Bos, Poephagus, Bibos, Bubalus, respectively. Divergency times among bovine species were between 0.775-6.43 MYA. The longest divergency time was between Asian water buffalo and other bovine species. The shortest divergency time was between Bos taurus and Bos indicus. The monophyletic relationship was not emerged between Bos frontalis and Bos gaurus, Bos frontalis and Bos gaurus did not differentiated clearly on mitochondrial DNA.【Conclusion】Six domestic Chinese Bovinae species could be divided into four branches corresponding to Bos, Poephagus, Bibos and Bubalus, respectively. The relationships between Bos and Poephagus, Bibos, Bubalus deduced in turn gradually. Divergency times among six bovine species were between 0.775-6.43 MYA. There was very close relationship between Bos frontalis and Bos gaurus, which showed that they might share the same maternal origin in an anterior time. The results did not support the opinion that Bos frontalis might be the domesticated form of Bos gaurus, Bos frontalis and Bos gaurus might be the progeny of a kind of extinct wild bovine species.

Effect of Maternal Feed Restriction on Serum Lipid Metabolism During Embryo Period

ZHANG Yuan-Yuan, DAN An-Shan, LI Feng, HU Jing-Wei, WANG Lian-Sheng, LI Yao-Hua, CHENG Bao-Jing, BI Zhong-Peng

Scientia Agricultura Sinica. 2011, 44(19): 4088-4095. doi:10.3864/j.issn.0578-1752.2011.19.020

Abstract ( 744 )

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【Objective】 An experiment was conducted to evaluate the effects of maternal feed restriction on the offspring embryo period serum biochemical indices and hormone levels in relation to lipid metabolism. 【Method】 Three hundred and eighty-four fat line and 384 lean line broiler breeders bred by Northeast Agricultural University were respectively divided into two groups. During the laying period, group one was fed normally, and the daily feed intake of group two was 75% of group one. The eggs were collected and hatched when the laying rate rose to the peak. After collecting 150 eggs according to each maternal group, blood samples were taken at 13 embryonic day (E), 15E, 17E and 19E. 【Result】 The results showed that the maternal feed restriction significantly decreased the concentration of CHO and TG in serum of 13 embryonic day (E) and LDL in serum of 15E (P＜0.05); While, the maternal feed restriction significantly increased the concentration of HDL in serum of 19E and G in low fat line of 13E and 15E (P＜0.05). The maternal feed restriction significantly decreased the concentration of GLU in serum of 15E high fat line, 19E and INS of 13E, 15E and 19E and LEP of 13E and 15E (P＜0.05). During four days, serum GH level of feed restriction group was generally lower than the normal feeding group. 【Conclusion】 The results demonstrated that maternal feed restriction on the laying period directly affected serum lipid metabolism in embryo period and the effect occurred in 13E. Different trends of serum biochemical indices were found in different breeders.

Co-expression of MIC-5 Gene of Eimeria stiedai and Rabbit IL-15 in Eucaryotic Cell Line

MENG Qing-Ling, QIAO Jun, CAI Xue-Peng, TIAN Guang-Fu, YAN Hong-Bin, LUO Xue-Nong

Scientia Agricultura Sinica. 2011, 44(19): 4096-4101. doi:10.3864/j.issn.0578-1752.2011.19.021

Abstract ( 520 )

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【Objective】 The aim of this study is to construct a recombinant co-expression vector expressing Eimeria stiedai microneme protein 5 (MIC-5) and rabbit IL-15 in eukaryotic cells, which may lay a foundation for further study immune regulatory DNA vaccine against infection of Eimeria stiedai. 【Method】 E. stiedai MIC-5 gene and rabbit's IL-15 gene were respectively cloned into plasmid pBudce4.1 to construct recombinant vector pB-MIC-5-IL-15 for co-expression, and then the recombinant plasmid pB-MIC-5-IL-15 was transfected into BHK-21 cell line for expression. The RT-PCR and indirect fluorescent immunoassay were used to detect the transcription and expression of interesting genes in BHK-21 cell line, respectively. 【Result】 The results showed that E. stiedai MIC-5 gene and rabbit's IL-15 gene were successfully transcripted. And interesting genes of E.stiedai MIC-5 and rabbit's IL-15 were successfully expressed at 36, 48 and 72 hour post-transfection, respectively. 【Conclusion】 The recombinant co-expression vector was successfully constructed, which can express E. stiedai MIC-5 and rabbit IL-15 gene in eukaryotic cells.

Studies on Pharmacodynamics in vitro of the Florfenicol Dual Suspension Emulsion

LI Xiao-Wen, DONG Wei, YANG Kun, CHEN Xiao-Jun, LIU Zhao-Ying, SUN Zhi-Liang

Scientia Agricultura Sinica. 2011, 44(19): 4102-4109. doi:10.3864/j.issn.0578-1752.2011.19.022

Abstract ( 776 )

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【Objective】 The in vitro pharmacodynamics characteristics of florfenicol dual suspension emulsion (DSEF) were studied in an experiment. 【Method】 The florfenicol injection (FI) was used as the control group and five kinds of bacteria were used for in vitro antibacterial test to determine the minimal inhibitory concentration (MIC), minimal bactericide concentration (MBC) and mutant selection window (MSW), respectively. The S.typhimurium was used for in vitro antibacterial test about post-antibiotic effect (PAE) and post-antibiotic sub-MIC effect (PASME), respectively. 【Result】 Compared with the ordinary injection, the MIC and MBC of DSEF for five kinds of bacteria showed no change obviously. But DSEF narrowed MSW obviously of five kinds of bacteria (P＜0.01) and extended the PAE and PASME significantly of S. typhimurium (P＜0.01). 【Conclusion】DSEF could reduce the bacterial resistance risk and extend the time of its after effect obviously, thereby improved the antibacterial effect.

Study on Comprehensive Evaluation and Influencing Factors of Information Service in Rural Areas——Based on the Survey of Village-Level Perspective in Ningxia Hui Autonomous Region

LI Jin, ZHAO Chun-Jiang, QIN Xiang-Yang, YUAN Xue-Guo, ZHANG Zheng

Scientia Agricultura Sinica. 2011, 44(19): 4110-4120. doi:10.3864/j.issn.0578-1752.2011.19.023

Abstract ( 727 )

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【Objective】 The establishment of rural information service system is a systematic engineering which is influenced by various elements such as national conditions, regional characteristics, rural economic and social development. In order to understand the status quo，the effect and key influencing factors of rural information service, address the key working and then explore a mechanism of sustainable development for rural information service in new rural construction, this paper gives an analysis of the effect evaluation and influencing factors of rural information service patterns in Ningxia Hui Autonomous Region.【Method】The paper constructed a comprehensive evaluation index system for rural information service, combined with AHP and Delphi method to determine the index weight, then measured and evaluated it with Liken Scales. And what is more, it selected four factors for a fitting analysis like GDP, farmers’ culture quality, informatization input, net income per capita, network home rate, amount of information technology equipment per capita and information technology training person times seven factors of rural information service with the multiple linear regression model. 【Result】 The general level of information service was good from the sampled villages, farmers were satisfied with the content of information service and policy and legal guarantee. However, the evaluation on information capacities and construction of information resource were general according to these farmers. Rural information service was mainly relative to farmers culture quality, information input, amount of information technology equipment per capita and information technology training person times rather than GDP, net income per capita and network home rate. The more information input, the stronger the ability of farmers to accept the information, the better service effect. The more poverty backward area, the lower farmers’ culture quality got, the better effect of information service.【Conclusion】It was found that optimizing development environment was a key to solve “Last One Kilometer” of information service in rural area, information service can further expand its functions and effects in poorer regions, and moreover diversifying channels for information dissemination would greatly enhance its effects. Government should fully play a leading role in construction of rural information service system, optimize development environment of information service and speed up the popularization of portable information terminals, meanwhile, farmers’ consciousness and consumption level of information were needed to be improved .

Influence of Nitrate Nitrogen on Salt Tolerance and Osmoticum of Chinese Iris

ZHANG Biao, LI Pin-Fang

Scientia Agricultura Sinica. 2011, 44(19): 4121-4128. doi:10.3864/j.issn.0578-1752.2011.19.024

Abstract ( 662 )

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【Objective】The objective of this study was to investigate the salt tolerance of Chinese Iris (Iris lactea Pall. var. chinensis (Fisch.) Koidz.) and the osmoticum in leaves at different levels of exogenous nitrate, to provide a scientific basis for cultivation and management of Iris lactea. 【Method】 Seedlings in sand culture experiment were treated with nine cross combinations of different NaCl concentrations at 0.1, 140, and 210 mmol?L-1, and three nitrate nitrogen levels at 0.25, 4 and 8 mmol?L-1, respectively. After 35 d of treatment, biomass of leaf and root, uptake of mineral elements and the main osmoticum in leaves were determined.【Result】NO3- at 8 mmol?L-1 caused a much higher increase in leaf biomass than 4 mmol?L-1 nitrate at 140 mmol?L-1 NaCl treatment. The exogenous nitrate (4 and 8 mmol?L-1) caused a 30% increase in leaf biomass at 210 mmol?L-1 NaCl, compared to the values at 0.1 mmol?L-1. No differences in leaf biomass were observed between two nitrate concentrations (P＜0.05). Exogenous nitrate resulted in a decrease in root/shoot ratio and leaf membrane permeability but an increase in total N of leaf and root under NaCl stress, while it had no significant effect on mineral element contents (K+、Na+、Ca2+、Mg2+). Contents of inorganic ions in leaves under interactions of nitrate and NaCl was Cl-＞K+＞Na+＞NO3-. Increase in concentrations of exogenous nitrate caused a decrease in Cl-, Na+ and K+ contents but an increase in proline and NO3- contents.【Conclusion】The growth under NaCl stress was improved by exogenous medium concentration of nitrate, and the mechanism may depend on improvement of nutrient uptake under NaCl stress, increase in proline and NO3- and decrease in root/shoot ratio, Cl-, Na+ and K+ contents.

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