Scientia Agricultura Sinica

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Scientia Agricultura Sinica. 2020, 53(7): 0.

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Research Progress of Wheat Wild Hybridization, Disease Resistance Genes Transfer and Utilization

Cheng LIU,Ran HAN,XiaoLu WANG,WenPing GONG,DunGong CHENG,XinYou CAO,AiFeng LIU,HaoSheng LI,JianJun LIU

Scientia Agricultura Sinica. 2020, 53(7): 1287-1308. doi:10.3864/j.issn.0578-1752.2020.07.001

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Wheat alien species are vast reservoir of diversity for disease and pest resistance as well as stress tolerance, which are excellent gene sources for wheat breeding. Through wide hybridization, the genes of alien species could be transferred to wheat to create wheat-alien chromosome lines such as amphiploids or partial amphiploids, additions, substitutions and translocation lines. These genetic stocks could be utilized to study chromosome behavior and genome evolution, mapping genes, and diversifying the genetic basis of wheat for diseases and pest resistance, as well as yield and quality improvement. In order to understand the progress of wheat wide hybridization and useful gene transfer from alien species to wheat, in this paper, the classification of the tribe Triticeae, the definition and significance of wheat wide hybridization, alien transfers progress from species belonging to genera Aegilops, Secale, Thinopyrum, Dasypyrum, Agropyron, Hordeum, Elymus, Leymus, Psathyrostachys and Eremopyrum to wheat have been summarized and discussed. To date, the official designated genes originated from wheat alien species include 17 stripe rust resistance genes, 35 leaf rust resistance gens, 30 stem rust resistance genes, 41 powdery mildew resistance genes, 3 Fusarium head blight-resistance genes, one wheat blast resistance gene, one Septoria tritici blotch resistance genes, one Septoria nodorum blotch resistance gene, 4 tan spot resistance genes, 2 eyespot resistance genes, one wheat spindle streak mosaic virus resistance gene, 2 wheat streak mosaic virus resistance genes and 2 cereal yellow dwarf resistance genes. Names and the chromosomal locations of these disease resistance genes were inducted. Moreover, the utilization of these genes in wheat breeding has also been reviewed and summarized. In the history of world wheat breeding, disease resistant germplasms such as wheat-rye 1RS·1BL translocation, 1RS·1AL translocation and wheat-Aegilops ventricosa 2NS/2AS translocation have made outstanding contributions. However, this only benefited from the utilization of a few disease resistant genes. Compared to the number of the designated genes, relatively few disease-resistant genes have been used in wheat breeding. In this paper, the limiting factors for the underutilization are discussed. Suggestions on how to use these disease-resistant genes in the future are put forward. Meanwhile, the cloned disease-resistant genes from wheat alien species are listed. The methods of cloning these genes and the possible research hotspots in the future are also analyzed. It is believed that the development and application of wheat-wild species translocation lines without genetic drag may be an important driving force for material innovation and variety breeding in the future.

Analysis of Quality Traits and Breeding Inspiration in Yangmai Series Wheat Varieties

Xiao ZHANG,Man LI,DaTong LIU,Wei JIANG,Yong ZHANG,DeRong GAO

Scientia Agricultura Sinica. 2020, 53(7): 1309-1321. doi:10.3864/j.issn.0578-1752.2020.07.002

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【Objective】 The objectives of this study were to evaluate the quality traits of Yangmai series wheat varieties and carry out quality classification , thus providing guidance for planting appropriate wheat varieties in the suitable areas and selecting proper parents for wheat quality breeding. Moreover, based on the analyses between the performances of quality traits and pedigree of different types of varieties, a simple and practical quality selection index and principle of parent selection were proposed. 【Method】 A total of twenty-four Yangmai wheat varieties were planted in randomized complete blocks with two replications at the research station of Lixiahe Institute of Agricultural Sciences of Jiangsu during the 2016 and 2017 cropping seasons. Quality parameters, such as kernel hardness, grain protein content, wet gluten content, SDS sedimentation value, solvent retention capacity (SRC), farinograph parameters and rapid visco analyzer parameters were investigated after harvesting. 【Result】 The hardness of tested wheat varieties were ranged from 13.48 to 62.12, those can be divided into soft wheat (lower than 32) and hard wheat (higher than 54); In addition, the values of sedimentation value, protein content, wet gluten content, gluten index, water SRC, sodium carbonate SRC, sucrose SRC, absorption, development time, stability time, peak viscosity and pasting temperature were 6.33-13.75 mL, 12.60%-14.61%, 29.74%-38.13%, 38.86%-82.83%, 54.69%-78.56%, 71.40%-107.73%, 95.66%-127.27%, 54.77%-67.40%, 1.23-8.83 min, 2.20-13.17 min, 3 000 cP up and down, and 61.57-64.75℃, respectively. We found that Yangmai4, Yangmai10, Yangmai158, Yangmai16, Yangmai17 and Yangmai23 showed significantly higher hardness, water SRC, sodium carbonate SRC, sucrose SRC and water absorption than other varieties based on the analysis. However, we did not find the significant different of protein content and wet gluten content between different varieties. Development time and stability time of Yangmai 2 was the longest, wherea no significant differences were observed among most varieties. The cluster analysis indicated that 24 varieties could be categorized into two major groups, which was in consistence with the results from pedigree analysis. 【Conclusion】 Most of Yangmai series wheat varieties could be classified as weak gluten wheat with low hardness, gluten index, water SRC, sodium carbonate SRC, sucrose SRC and water absorption. However, Yangmai4, Yangmai10, Yangmai158, Yangmai16, Yangmai17, and Yangmai23 were mid-strong gluten wheat with relatively high hardness, gluten index, water SRC, sodium carbonate SRC, sucrose SRC and water absorption. Yangmai series wheat varieties generally showed high peak viscosity and low pasting temperature. Hardness can be used as an efficient and practical selection index for quality breeding. We recommend that wheat breeders have to use at least one corresponding quality cultivar/line as one of parent for making cross if you are going to develop good quality wheat variety.

Development and Application of Plasmid Reference Molecule for Genetically Modified Rapeseed Screening

Jun LI,Xia-ying LI,Jing-qian WANG,Shanshan Zhai,Zi-yan CHEN,Hong-fei GAO,YunJing LI,Gang WU,Xiu-jie ZHANG,Yu-hua WU

Scientia Agricultura Sinica. 2020, 53(7): 1322-1337. doi:10.3864/j.issn.0578-1752.2020.07.003

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【Objective】 Rapeseed is one of the four major genetically modified (GM) crops, the production and application of GM rapeseed must be regulated in China. Performance of GMO detection is the prerequisite to implement GMO regulations, screening is the first step to determine the presence or absence of GMO ingredients in testing samples. Appropriate selection of screening targets can effectively reduce the chance of missed detection of some GM ingredients. A technical platform, involving establishment of screening strategy for GM rapeseed and development of a common reference plasmid that is compatible with the screening strategy, would provide technical support for regulating GM rapeseed.【Method】 Both regulatory elements and marker genes commonly used in GM rapeseed are obtained by collecting and analyzing the GM rapeseed varieties registered in database, then the screening strategy for GM rapeseed can be determined based on the principle of maximum coverage of GM rapeseed varieties. The whole nucleotide sequences of screening elements are collected by searching nucleotide database or retrieving patent. One screening target usually has multiple standard detection methods, both the primer pairs for conventional PCR and the primers/probe combinations for real-time PCR are aligned with the nucleotide sequence of each screening target to determine the target sequence that would be integrated into plasmid. The fusion sequence of all screening elements together with rapeseed reference genes was artificially synthesized, and cloned into the plasmid pUC18 to construct a positive plasmid molecule. Both conventional PCR and real-time PCR are utilized to evaluate the applicability of constructed plasmid as positive control.【Result】 The screening strategy for transgenic rapeseed was established by detecting nine elements, involving two promoters of CaMV 35S and FMV 35S, five genes of Bar, PAT, CP4-EPSPS, NPTII, and HPT, two terminators of NOS and PinII. This screening strategy achieved full coverage of transgenic rapeseed varieties with known information. The screening plasmid pYCSC-1905 was constructed for GM rapeseed, carrying nine screening elements and 2 rapeseed reference genes of HMG I/Y and CruA. The amplification efficiencies of nine screening elements and two rapeseed reference genes were in the range from 90% to 110%, demonstrating that the amplification efficiency of screening target is not influenced due to the mutual interference of integrated fragments. The plasmid pYCSC-1905 can be used as a common positive control for nine screening targets and two rapeseed reference genes, applicable to national standards (GB/T and Declaration of Ministry of Agriculture and Rural Affairs), Entry-exit inspection and quarantine industry standards (SN/T) and European Union standards.【Conclusion】 The screening strategy covering 9 elements for GM rapeseed screening , can achieve the screening of GM rapeseed in all stages from commercialization to safety assessment, and significantly reduce the missed detection of GM rapeseed. The developed plasmid pYCSC-1905 provides a general positive control for rapeseed screening and the standard methods, and ensures the accuracy and comparability of test results between laboratories.

The Impacts of Post-Anthesis Warming on Grain Yield and Quality of Late Japonica Rice in a Double Rice Cropping System

TaoTao YANG,JiaXin XIE,Shan HUANG,XueMing TAN,XiaoHua PAN,YongJun ZENG,QingHua SHI,Jun ZHANG,YanHua ZENG

Scientia Agricultura Sinica. 2020, 53(7): 1338-1347. doi:10.3864/j.issn.0578-1752.2020.07.004

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【Objective】 The aim of this study was to evaluate the response of the grain yield and quality of late japonica rice in a double rice cropping system to post-anthesis warming, so as to provide a theoretical basis for the safe production and high quality cultivation of double-cropped late japonica rice under the future warming conditions.【Method】 This experiment was conducted at Shanggao Experimental Station of Jiangxi Agricultural University in 2017 and 2018, which was located in the main producing area of a double rice cropping system. The field warming experiment consisted of post anthesis warming treatment (PAW: increases temperature day and night continuously from heading stage to maturity stage) and ambient temperature treatment (CK) with three replicates in a randomized complete block design. The tested rice cultivar was indica-japonica hybrid rice Yongyou1538. PAW treatment was treated with a free-air temperature increase (FATI) facility. Grain yield, yield components, milling quality, appearance quality, RVA, amylose content, protein content, and amino acid content were compared and analyzed between control and post-anthesis warming treatments. 【Result】 Grain yield and quality of Yongyou 1538 changed obviously under PAW, and the change trend was basically consistent in two years. The specific performances were as follows: compared with CK, post-anthesis warming with an average canopy temperature increase of 2.2℃ decreased the grain yield by an average of 4.4% in two years. PAW had no significant effects on the milling quality, but increased chalky grain rate and chalkiness by an average of 27.4% and 24.4% in two years, respectively. Peak viscosity, trough viscosity, and pasting temperature showed increase trends under post-anthesis warming condition, but decreased setback, and had no effects on breakdown and final viscosity. The amylose content was decreased by an average of 6.4%, but the protein relative and absolute contents in milled rice were increased by an average of 8.7% and 6.6%, respectively. In addition, the content of non-essential amino acids and essential amino acids in milled rice were both increased under PAW, and the response of non-essential amino acids to PAW was more sensitive than that of essential amino acids, among which the total amount of non-essential amino acids was significantly increased by 5.7%. From the relative ratio of amino acids, it could be found that PAW had no significant effect on the relative ratio of most amino acids, but Tyr and Met. 【Conclusion】 Post-anthesis warming decreased grain yield of late japonica rice, deteriorated its appearance and cooking quality, but benefited its nutritional and eating quality.

Effects of Climatic Factors Under Diverse Ecological Conditions on Foxtail Millet (Setaria italica) Yield in Shandong

YanBing YANG,Ling QIN,RunFeng WANG,ErYing CHEN,XiuBo YIN,YuQin LIU,SuMei ZHANG,XinJun CONG,GuoYu LI,LeZheng WANG,YanAn GUAN

Scientia Agricultura Sinica. 2020, 53(7): 1348-1358. doi:10.3864/j.issn.0578-1752.2020.07.005

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【Objective】Grain yield is one of the most important indicators in evaluating foxtail millet productivity and is greatly affected by the fluctuation of climatic factors. The researches on the effects of climatic factors on grain yield and the varietal response of grain yield to different ecological conditions in Shandong can provide scientific basis for selecting, breeding and planting high-quality foxtail millet. 【Method】 Eight newly released foxtail millet cultivars selected from the summer-sown region of North China were planted at five locations in two consecutive years (2016 and 2017). Grain yields of these cultivars were measured. Analysis of Variance (ANOVA) was used to determine the significance of differences in grain yields by Duncan’s Multiple Range Test. And correlation analysis between grain yield and air temperature, rainfall, and sunshine duration were conducted by Pearson’s method. 【Result】 ANOVA showed that growing years, growing locations × growing years, and growing locations × cultivar genotypes had highly significant effects on the variations in grain yields (P<0.01). Similarly, cultivar genotypes, cultivar genotypes × growing years, and growing years × cultivar genotypes × growing years showed significant influence on grain yields (P<0.05). Growing locations, growing locations × growing years, growing locations × cultivar genotypes and cultivar genotypes had high contribution rates (50.05%, 19.76%, 12.32%, and 8.67%, respectively), whereas growing years, cultivar genotypes × growing years had low contribution rates (2.70% and 1.69%, respectively) to the variations in grain yields. In 2017, the average yield per unit area of eight foxtail millet cultivars was 4.55% higher than that in 2016, however, that was not consistent between growing locations. The annual outputs from Jinan, Linyi and Jining in 2017 were increased by 21.64%, 18.47% and 3.96%, respectively, while from Dezhou and Taian the annual outputs were decreased by 8.66% and 9.78%, respectively, compared with those in 2016. The average yields of eight cultivars from five locations across two years were 5 657.2 kg·hm^-2 ranging from 5 267.8 kg·hm^-2to 5 926.0 kg·hm^-2. The yield of Yugu18 had the lowest yield while Jigu20 had the highest yield. No significant difference in grain yields was observed between cultivars Jigu20, Jigu22, Jilügu1, Jinuogu2 and Zhonggu2, however, grain yields of these cultivars were significantly higher than Jigu19 and Yugu18. Correlation analysis showed that the grain yield was positively but not significantly correlated with the average temperature and maximum temperature at each time period (P>0.05). However, it was significantly positively correlated with the total average temperature of the whole growth period (P<0.05). On the other hand, grain yield was negatively correlated with the rainfall at seedling stage (middle and late June) (P>0.05), and the rainfall during the periods from pregnancy heading to flowering (from late July to middle August) (P>0.05). Furthermore, grain yield was positively correlated with sunshine duration in middle and late September (P>0.05). 【Conclusion】 Growing years, growing locations, cultivar genotypes, growing locations × growing years and growing locations × cultivar genotypes had significant effects on the yields of foxtail millet in Shandong. The fluctuation of ecological factors across different years and different locations, especially the air temperature and rainfall were the important factors affecting the yields. Higher average air temperature during the whole growth period was beneficial to increasing grain yield. Excessive rainfall at seedling and heading stages had negative effects on grain yields. The yield of testing cultivars was significantly different between years and between experimental sites. The cultivars, Jigu20, Jigu22, Jilügu1, Jinuogu2 and Zhonggu2 had good fertility and stability across ecological conditions.

Impacts of Decomposing Microorganism Inoculum on Straw Decomposition and Crop Yield in China: A Meta-Analysis

XinRun YANG,Bei XU,ZhiFeng HE,Jing WU,RuiHua ZHUANG,Chao MA,RuShan CHAI,Kianpoor Kalkhajeh Yusef,XinXin YE,Lin ZHU

Scientia Agricultura Sinica. 2020, 53(7): 1359-1367. doi:10.3864/j.issn.0578-1752.2020.07.006

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【Objective】 Whether the application of straw decomposing microorganism inoculum (SDMI) in cropland could significantly promote returning straw decomposition and their nutrients releasing, and thus improve crop yield is still on debate. Therefore, it is very necessary to meta-analysis the effects of the SDMI on returning straw decomposition and crop yield on a national scale, as well as explore the variation of the effect under different climates, straw characteristics and returning conditions providing guidance for SDMI application. 【Method】The meta-analysis was performed involving 63 comparisons of straw decomposition and 207 comparisons of crop yield from 89 studies during 2001 and 2009 to identify site-specific factors, including regional climates, returning conditions, straw types, initial returning carbon-to-nitrogen ratio (C/N), soil pH, and soil organic matter (SOM). 【Result】 The results showed that SDMI application could significantly increase the decomposition rate of returning straw, except the study was designed at temperate continental climate areas (range of effect size was from -0.010 to 0.716, the same in below), or initial returning C/N was no more than 15 (-0.302 to 0.568), or tested soil was neutral (95% CI of effect size was from -1.01 to 0.475). In addition, the strength of SDMI application could be varied significantly under different returning conditions, straw types, and SOM. Similar to straw decomposition, SDMI application also could significantly increase the crop yield, except the study designed at neutral soil (-0.284 to 0.751). Regional climates, straw types, initial returning C/N, soil pH rather than returning conditions and SOM would regulate the increase rate of crop yield. Moreover, the linear fitting indicated the positive correlation between the rate of straw decomposition and crop yield (R²= 0.307, P<0.0001). 【Conclusion】 This study revealed that SDMI application in Chinese farmland could significantly promote returning straw decomposition and crop yield increasing, especially in temperate monsoon climate area with dryland, maize straw returning, initial returning C/N≥30 and acidic or alkaline soil.

Biologic and Transcriptomic Analysis of Citrus hystrix Responses to ‘Candidatus Liberibacter asiaticus’ at Different Infection Stages

CaiLing TENG,Xi ZHONG,HaoDi WU,Yan HU,ChangYong ZHOU,XueFeng WANG

Scientia Agricultura Sinica. 2020, 53(7): 1368-1380. doi:10.3864/j.issn.0578-1752.2020.07.007

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【Objective】 Citrus Huanglongbing (HLB), associated with phloem-colonized ‘Candidatus Liberibacter asiaticus’ (CLas), severely impedes worldwide citrus production. The objective of this study is to analyze the biological symptoms, microstructures and transcriptomes of Citrus hystrix in response to CLas infection at different stages, to reveal the tolerance mechanism of C. hystrix, and to provide a basis for further screening of resistance genes and HLB-tolerant/resistant citrus breeding. 【Method】| The budwoods of C. hystrix grafted on two-year-old Carrizo citrange rootstocks (C. sinensis× P. trifoliata) used in this study were graft-inoculated with budwoods from a CLas (strain GZBJT)-infected Guanximiyou pummelo maintained in a greenhouse at Citrus Research Institute. The budwoods used as inoculum were tested CLas positive and free of other potential phloem-limited pathogens, such as Citrus tristeza virus (CTV) or Citrus tatter leaf virus (CTLV) by PCR before grafting. Inoculated plants were kept in greenhouse along with mock-inoculated healthy control plants. Real-time quantitative PCR (qPCR) was performed every 15 days after inoculation. Four months after inoculation (the earliest establishment of CLas by qPCR) and 14 months after inoculation were defined as the early infection stage and the late infection stage, respectively. The biological symptoms and microstructures were observed to analyze the structural changes of different infection stages. Combined with comparative transcriptome and RT-qPCR validation, the response mechanism of C. hystrix against HLB was explored.【Result】 No typical symptom was observed in C. hystrix at the early and late stages of infection. Light microscopy observation from the midribs of HLB-affected and uninfected C. hystrix revealed that no significant structure change was found at the early infection stage and only a few sieves in the phloem were blocked at the late stage. By comparing the RNA-seq data, 181 and 1 384 genes were found to be differentially expressed at the early stage and late stage, respectively. Differentially expressed genes (DEGs) mainly involved in cell wall metabolism, host defense response, starch and sucrose metabolism, callose synthesis and signal transduction. Comparative transcriptome analysis showed that the expression of related genes in starch and sucrose metabolism and cell wall metabolism was down-regulated at the early infection stage, and the expression of related genes in salicylic acid metabolism, salicylic acid signal transduction pathway, pathogenesis-related protein and glutathione-S-transferases was up-regulated at the late infection stage.【Conclusion】 The early response of C. hystrix to CLas infection is mainly characterized by stable physical structure, undisturbed pathways such as starch synthesis and photosynthesis. Salicylic acid-mediated resistance signals, effector-triggered immunity (ETI), and glutathione-S-transferases mediated detoxification contribute to the tolerance of C. hystrix against CLas at the late infection stage.

Differential Expression of LncRNAs in Nicotiana tabacum var. Samsun NN Infected by TMV at Different Temperatures

HaiYan JIA,LiYun SONG,Xiang XU,Yi XIE,ChaoQun ZHANG,TianBo LIU,CunXiao ZHAO,LiLi SHEN,Jie WANG,Ying LI,FengLong WANG,JinGuang YANG

Scientia Agricultura Sinica. 2020, 53(7): 1381-1396. doi:10.3864/j.issn.0578-1752.2020.07.008

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【Objective】 The objective of this study is to screen out the differentially expressed lncRNAs in Nicotiana tabacum var. Samsun NN after Tobacco mosaic virus (TMV) infection at different temperatures, and to investigate the role of identified lncRNAs in Samsun NN’s resistance response.【Method】 The temperature sensitivity of N gene makes Samsun NN have the resistance to TMV at 25℃, but lose it at 31℃. TMV and phosphate buffered saline (PBS) were mechanically inoculated into Samsun NN at 25℃ and 31℃. Total RNAs were extracted from systemic leaves at 48 hpi (hours post infection). Deep sequencing was performed after strand-specific database construction, and the sequencing results were filtered to get clean reads. Using N. tabacum var. TN90 as a reference, HTseq was employed to compare obtained reads. LncRNAs were screened out, and their expression levels were estimated by FPKM method. Differentially expressed lncRNAs (DElncRNAs) were then identified by edgeR and verified by qRT-PCR. The target genes of DElncRNAs were predicted by co-localization and co-expression analyses. Gene annotations, GO and KEGG pathways were analyzed for functional prediction of target genes. 【Result】 Altogether, 80 million clean reads were detected for each of 12 samples from 4 treatments by lncRNA-seq. A total of 4 737 annotated lncRNAs and 40 169 novel lncRNAs were obtained. Among them, 64 lncRNAs were differentially expressed after TMV infection at different temperatures. qRT-PCR results showed that the sequencing accuracy of these DElncRNAs was about 80%, which indicated the sequencing data obtained in this study had high reliability. Importantly, some target genes were simultaneously targeted by DElncRNAs that were down-regulated at 25℃ and up-regulated at 31℃. Gene annotations showed that DElncRNAs’ target genes involved in many functions, such as plant resistance, hormone and metabolic pathways. Particularly, some lncRNAs that may be associated with hormone pathways showed a down-regulation trend after TMV infection at 25℃, while up-regulated at 31℃. Furthermore, GO enrichment analysis showed that target genes were mainly involved in the composition of membranes, vesicles, and acted as calcium and potassium ion channel inhibitor activity, so that consequently the corresponding ions could be transported to their sites of action to trigger subsequent reactions. Moreover, these genes were also involved in pathogenesis, antigen processing and presentation, cytokinin metabolism and other physiological processes. The plant hormone signaling pathway was significantly enriched by target genes during KEGG pathway analysis. DElncRNAs related genes down-regulated at 25℃ and up-regulated at 31℃ were simultaneously enriched in pathways such as hormone signaling, ABC transporters, and phenylpropanoid biosynthesis.【Conclusion】 LncRNAs were differentially expressed in Samsun NN after TMV infection at different temperatures (25℃ and 31℃). DElncRNAs participated in host plant systemic acquired resistance by acting on hormone signaling transduction, substance transport and other processes. Taken together, this study lays a foundation for further research on lncRNA’s regulatory function in plant systemic acquired resistance and the development of new strategies to overcome virus invasion into host plant.

Quality Barrier Characteristics of Cultivated Layer for Sloping Farmland in Purple Hilly Region

Ge SONG,DongMei SHI,XiaoYing ZENG,GuangYi JIANG,Na JIANG,Qing YE

Scientia Agricultura Sinica. 2020, 53(7): 1397-1410. doi:10.3864/j.issn.0578-1752.2020.07.009

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【Objective】 The purple soil slope farmland is characterized by high productivity and strong erodibility, and is an important cultivated land resource in the upper reaches of the Yangtze River. Analyzing the types of barrier tillage layer of slope farmland and its impact on crop yield has important application value for determining reasonable tillage layer construction parameters, controlling tillage layer quality and sustainable utilization of slope farmland. Based on the sample of cultivated layer of purple soil slope farmland in different soil fertility grades, this paper quantitatively analyzed the quality barrier characteristics of slope farmland.【Method】 In this paper, the obstacle degree model and cluster analysis were used to study the types and quality characteristics of barrier tillage layer in purple soil slope farmland of different soil fertility grades.【Result】 The results showed that: (1) There were significant differences in soil physical properties between purple soil slope farmland with different soil fertility grades. With the decrease of soil fertility grade, the slope of the plot was significantly larger, and the thickness of the effective soil layer of plot was significantly smaller. When the slope of the plot changed from 5.1° to 21.7°, the crop yield could be reduced by 45%. The lack of soil layer in the five-slope farmland was serious. However, the soil fertility attributes did not show significant differences. The soil organic matter, soil total nitrogen and cation exchange capacity of the first to fourth grade slope farmland in the same soil fertility grade were as follows: topsoil layer>subsoil layer>bottom soil layer. There was no significant difference between the topsoil layer and the subsoil layer in the fifth grade slope farmland. (2) The soil quality index of each soil layer in the first and second grade slope farmland was between 0.434 and 0.528. The soil quality index of each soil layer of the same soil productivity grade was characterized by topsoil layer>subsoil layer>bottom soil layer; the soil physical properties had a more significant effect on the soil quality of low-yield slope farmland. The obstacle degree of the slope of the plot of the fifth-grade slope farmland was 80.73 times than that of the first-grade slope farmland. (3) The purple soil slope farmland could be classified into three types, namely, III soil nutrient limited type, IV effective soil layer thickness limited type, and V field surface slope limited type. When the soil physical property was the main obstacle feature, the cultivated layer structure was loose, the subsoil layer was seriously deficient, and the crop yield was 25% lower than that of the soil nutrient-restricted sloping farmland. 【Conclusion】 The quality of the cultivated layer in the purple soil slope farmland was low, and the barrier cultivated layer included three types: soil nutrient limitation type, effective soil layer thickness limitation type, and slope of the plot limitation type. The physical properties of soil were the main limiting factors for the quality of the cultivated layer of purple soil slope farmland. The slope of the field was too large, and the thickness of the effective soil layer was shallow and thin.

Seasonal Variation and Differences of Microbial Biomass Phosphorus in Paddy Soils Under Long-Term Application of Phosphorus Fertilizer

Kai LIU,Jia LIU,XiaoFen CHEN,WeiTao LI,ChunYu JIANG,Meng WU,JianBo FAN,ZhongPei LI,Ming LIU

Scientia Agricultura Sinica. 2020, 53(7): 1411-1418. doi:10.3864/j.issn.0578-1752.2020.07.010

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【Objective】 The seasonal dynamics of microbial biomass phosphorus (MBP) in paddy soils under long-term different fertilization treatments were studied to reveal the relationship between the variation characteristics and soil phosphorus supply. 【Method】 Taking the long-term experimental plot as a platform, four different fertilization treatments, including CK, NK, NP and NPK, were set up to collect 0-20 cm soil layer at tillering stage, booting stage, filling stage and full ripe stage of rice, respectively, and to determine soil total phosphorus (TP), available phosphorus (AP), acid phosphatase (ACP) activity and MBP.【Result】 Compared with CK and NK treatments, NP and NPK treatments significantly increased the contents of TP and AP in soil, with an increase of 88%-118% and 337%-903%, respectively. MBP was significantly affected by fertilization treatment, and was significantly higher in phosphate fertilization treatment than in non-phosphate fertilization treatment, NP and NPK treatments increased 103%-250% in the whole growth period (except tillering stage) compared with CK and NK treatments; seasonal variation of MBP showed a trend of first increasing and then decreasing and reached the highest level in the filling stage of rice in all fertilization treatments. The activity of acid phosphatase in NK treatment was the highest at grain filling stage, 38% higher than that under CK. At the same time, the turnover rate of microbial biomass phosphorus under this treatment was the highest, which could be recycled 1.31 times in the whole growth period. The correlation analysis showed that soil microbial biomass P was positively correlated with soil total P and available P. 【Conclusion】 The variation of microbial biomass phosphorus with different fertilization treatments and rice growth period was obvious, which was closely related to soil phosphorus availability.

Characteristics and Influencing Factors of Phosphorus Flows in the Crop-Livestock System of Fujian Province from 1985 to 2015

DongHui LIU,ShiChang ZHANG,Jing YANG,MengYuan HUANG,LiangQuan WU

Scientia Agricultura Sinica. 2020, 53(7): 1419-1431. doi:10.3864/j.issn.0578-1752.2020.07.011

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【Objective】The objective of this study is to determine the temporal changes and influencing factors of phosphorus (P) flows in the crop-livestock system of Fujian Province, and to provide a scientific reference for integrated nutrient management and green development of agriculture.【Method】Using a coupled NUFER (nutrient flows in food chain, environment and resources use) model, data were collected and sorted from statistical yearbooks, literatures, and field investigations from 1985 to 2015 to quantitatively estimate the balance, use efficiency and loss characteristics of P flow in the crop-livestock system of Fujian Province. 【Result】The total P input in crop-livestock production system increased from 63.1 to 196.2 Gg in Fujian Province from 1985 to 2015. The main P format inputs were fertilizer and feed import. The amount of per unit area of P fertilizer input in crop system increased gradually from 27.8 to 60.4 kg·hm^-2. Due to the influence of scale and structure change of crop-livestock system in Fujian Province, local feed supply decreased from 3.33 to 1.65 Gg, while the feed import increased from 20.7 to 70.2 Gg. From the output perspective, total P increased from 45.0 to 90.9 Gg, mainly including crop main product and animal manure loss. P uptake of crop main products changed slightly (only from 24.3 to 26.7 Gg), while the P loss in animal manure increased significantly from 1.44 to 25.8 Gg. Soil accumulation is the main loss pathway of P in crop-livestock system, which increased year by year from 18.1 to 106 Gg. P use efficiency in crop production system (PUE_c) decreased from 36.1% to 16.6%, and the change trend of P use efficiency in crop-livestock system (PUE_c+a) was similar to that of PUE_c, which gradually decreased and eventually remained at 15.0%. In the same time, the unit P loss of crop-livestock products increased gradually from 0.3 to 1.3 kg P·kg^-1. In terms of economic development and planting structure, there was a significant positive correlation between per capita GDP and fertilizer input when the per capita GDP was less than 11 000 yuan. When per capita GDP was less than 15 000 yuan, there was a significant positive correlation between per capita GDP and unit P loss of crop-livestock products, but there was a significantly negative correlation between per capita GDP and PUE_c when per capita GDP was less than 13 000 yuan. The proportion of cash crops planting area was significantly and positively correlated with the unit P loss of crop-livestock products and P input of fertilizer, and it was significantly negatively correlated with PUE_c.【Conclusion】At present, the cash crops planting area is relatively high, which is one of the structural characteristics of crop system in Fujian Province. At the same time, the unit area quantity of P input by fertilizer is large, circulation of animal manure is inadequate, which probably lead to more unit P loss in crop-livestock products, and low P use efficiency. Therefore, controlling the application amount of phosphate fertilizer and enhancing the utilization level in crop-livestock waste resources will be the guarantee in improving P use efficiency and promoting agriculture sustainable green development in Fujian Province.

Effects of Lower Limit of Drip Irrigation on Growth, Yield and Root Distribution of Greenhouse Grapes

Bo LI,Jun SUN,XinGuang WEI,SiYu ZHENG,Dong GE,ShiNing FU

Scientia Agricultura Sinica. 2020, 53(7): 1432-1443. doi:10.3864/j.issn.0578-1752.2020.07.012

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【Objective】 The experiment was carried out to explore the effects of irrigation level on grapes growth and water consumption under a controlled automatic irrigation, so as to provide a basis for grapes water management under an automatic irrigation in a greenhouse.【Method】 With the 3-year-old ‘Muscat Hamburg’ grape as the study object, a combined utilization of data acquisition terminal of CR1000, soil moisture sensor and radiotube, the experiment was carried out and set 8 different irrigation limits (accounting for 50%, 55%, 60%, 65%, 70%, 75%, 80% and 85% of field capacity, respectively), the same upper limit of irrigation being 90% of field capacity, so as to analyze the effects of different irrigation lower limits on biomass, yield and water consumption of ground and underground grapes in a greenhouse.【Result】 The results showed that the length and stem diameter of new shoots, as well as the leaf area index increased significantly with the increase of irrigation lower limits when it was lower than 75% of field capacity. The growth of new shoots was restricted to a certain degree when the lower irrigation limit exceeded 75% of field capacity. The grapes roots were distributed in soil layers of 0-60 cm, and the main distribution area was in 0-30 cm, where the volume and surface area of roots accounted for 75%-89% and 77%-83% of the total, respectively. In the most concentrated root distribution region in soil layer of 0-10 cm and 10-20 cm, there was a tendency to decrease after an increase in root index with the increase of lower irrigation limits; the root index reached the maximum with lower irrigation limit being 75% of field capacity. The root index increased with the increase of irrigation volume when the volume was lower than 6 000 m ³·hm ^-2, and decreased or increased slowly when the irrigation volume exceeded 7 000 m ³·hm ^-2. Grapes yield and water utilization efficiency reached the maximum with the lower irrigation limit being 75% of field capacity, which were 32 270.31 kg·hm ^-2 and 4.85 kg·m ^-3, respectively.【Conclusion】 Taking the factors such as new shoots growth, root distribution, grapes yield and water utilization into consideration, we could conclude that the optimal range of soil moisture was got when the lower irrigation limit took up 75% to 90% of water capacity under drip irrigation, which could be used as the recommended value of suitable irrigation control index under the planting mode.

Effects of Heat Stress on Physiological Indexes and Ultrastructure of Grapevines

Min LIU,Yulin FANG

Scientia Agricultura Sinica. 2020, 53(7): 1444-1458. doi:10.3864/j.issn.0578-1752.2020.07.013

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【Objective】 The objective of this research was to study the effects of heat stress on physiological indexes and ultrastructure of grapevines, and to explore the physiological response mechanism of grapevines to heat stress. 【Method】 One-year cutting seedlings of Vitis vinifera L. cv. Cabernet Sauvignon and Vitis davidii Foex. cv. Junzi 1were used as test materials, which were treated at 35℃, 40℃ and 45℃ for 48 h, with the plants at 25℃ as the control. Relative electrolyte leakage, relative water content, total chlorophyll content, antioxidant enzyme activity, malondialdehyde/super oxygen anion/hydrogen peroxide content, chlorophyll fluorescence and photosynthetic parameters of grape leaves were detected. By the scanning electron microscopy (SEM) and transmission electron microscopy (TEM), stomatal morphology and chloroplast ultrastructure were observed. 【Result】 After heat treatment at 35℃ and 40℃, the plant morphology, relative electrolyte leakage, relative water content, total chlorophyll content and chlorophyll fluorescence of Cabernet Sauvignon and Junzi 1 showed no significant changes, but net photosynthetic rate (Pn), transpiration rate (Tr) and stomatal conductance (Gs) were significantly reduced. After heat treatment at 45℃, Cabernet Sauvignon and Junzi 1 appeared stress symptoms, with relative electrolyte leakage increasing, and relative water content, total chlorophyll content, Fv/Fm, ΦPSII, Pn, Tr and Gs decreasing. The change range of each parameter in Junzi 1 was greater than that in Cabernet Sauvignon. With the increase of temperature, malondialdehyde/superoxide anion/hydrogen peroxide content, SOD and POD activities in grape leaves increased continuously, while CAT activity was the highest under moderate heat stress. The increase of SOD and CAT activities in Cabernet Sauvignon leaves was greater than that in Junzi 1. By SEM, it was found that at room temperature, the stomatal density of Cabernet Sauvignon was higher than that of Junzi 1; after heat treatment, the stomatal opening of grape leaves decreased, but the stomatal size did not change significantly. By TEM, it was found that after heat treatment, the chloroplast became large and round; the chloroplast membrane disintegrated; many giant starch grains appeared; a large number of plastoglobules were observed in the chloroplast of Junzi 1. 【Conclusion】 In Cabernet Sauvignon, stomatal density was high, chloroplast structure was stable, and SOD and CAT activities highly increased after heat treatment, which were the important reasons for its high heat resistance.

Phenolic Content, Bioavailability and Antioxidant Activity of Carambola

MuKang LUO,XuChao JIA,RuiFen ZHANG,Lei LIU,LiHong DONG,JianWei CHI,YaJuan BAI,MingWei ZHANG

Scientia Agricultura Sinica. 2020, 53(7): 1459-1472. doi:10.3864/j.issn.0578-1752.2020.07.014

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【Objective】 The aim of this study was to explore the phenolic composition, content, bioaccessibility and antioxidant activity of phenolic compounds in different carambola fruit varieties, so as to provide reference for the fresh consumption and deep processing of carambola. 【Method】 Phenolic compounds of three cultivars, including Guangzhou Hong cultivar (GZ), Xiangmi cultivar (XM), and Taiwan Misi cultivar (TW), were determined by high performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS). Their antioxidant capacity was evaluated by ABTS ⁺ radical scavenging capacity (ABTS) and oxygen radical absorbance capacity assay (ORAC). The change of total phenol, flavonoids, mono-phenol and their antioxidant capacity, as well as their bioaccessibility were analyzed during in vitro simulated gastrointestinal digestion. 【Result】 Three phenolic acids (protocatechuic acid, p-coumaric acid and ferulic acid) and three flavonoids (procyanidin B2, epicatechin and isoquercetin) were detected from the extracts of carambola fruit, among which proanthocyanin B2 and epicatechin were the predominant phenolics. The variation ranges of total phenolics and total flavonoids in tested varieties were 234.41-293.30 mg GAE/100 g and 165.75-278.97 mg CE/100 g, respectively. After simulated gastric digestion, the contents of phenolics and flavonoids in carambola digestive juice were 151.57-180.45 mg GAE/100 g FW and 113.06-164.45 mg CE/100 g FW, respectively. And their bioaccessibility were 54.05%-76.98% and 47.58%- 93.88%, respectively. After further intestinal digestion, the content of phenolic compounds released into the digestive fluid was reduced. The phenolics and flavonoids contents were 116.64-155.76 mg GAE/100 g FW and 78.47-148.44 mg CE/100 g FW, respectively, and their bioaccessibility were 46.57%-66.45% and 40.12%-84.75%, respectively. The contents of procyanidins B2 and epicatechin released into digestive fluid were 56.60%-87.54% and 65.33%-85.92% with gastrointestinal digestion and these only accounted for 51.90%-80.94% and 32.81%-37.50% of carambola fruit extract, respectively. The antioxidant capacity of ABTS and ORAC of digestive juice of three carambola fruit with gastric digestion were 97.56%, 102.42%, 92.36% and 122.73%, 118.50% and 107.14%, respectively. After further intestinal digestion, their antioxidant capacity of ABTS and ORAC were reduced by 12.33%-26.60% and 37.95%-43.28%, respectively. 【Conclusion】 Protocatechuic acid, p-coumaric acid, ferulic acid, procyanidin B2, epicatechin and isoquercetin were the main phenolic substances in carambola, and the contents of procyanidin B2 and epicatechin were the highest among them. After simulated gastric digestion, a high content of phenolic compounds in the digestive juice were detected, while their contents were significantly reduced with intestinal digestion.

Improving the Sensitivity of ELISA by Large-Capacity Reaction System of Aflatoxigenic Fungi-Biomarker in Agro-products

Xiao WEI,Qi ZHANG,Wen ZHANG,Hui LI,PeiWu LI

Scientia Agricultura Sinica. 2020, 53(7): 1473-1481. doi:10.3864/j.issn.0578-1752.2020.07.015

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【Objective】In order to prevent and reduce the contamination by Aspergillus species from the source, a highly sensitive large-capacity reaction system DAS-ELISA was established, based on the biomarker PO8 protein of aflatoxigenic fungi. This study aimed to provide key technical support for pollution source monitoring. 【Method】 In this study, the dry mycelium was used as a reference for the biomarker PO8 protein, the mycelia lysate made by high pressure homogenization was used as envelope antigen, the purified PO8-VHH was used as capture antibody, and rabbit polyclonal antibody against Aspergillus was used as detection antibody. The antigen and antibody were added to the 96-well microtiter plate at 200 μL/well, and the sandwich ELISA for the large-capacity reaction system was carried out. Based on the principle that the positive hole OD_450nm≥1.0, the positive hole OD_450nm/negative hole OD_450nm was higher to determine the optimal experimental conditions and to establish a standard curve. The performance of the established sandwich ELISA method was evaluated by spike-and-recovery test, repeatability test and specific test. 【Result】 Assays were performed in the PO8-VHH (3 μg?mL ^-1) coated ELISA format, in which the detection antibody was 2.5 μg?mL ^-1 diluted. The optimized physicochemical factors in the performance were obtained: the antibody coating condition was 4℃ overnight, the blocking reagent was 3% BSA, the blocking condition was 37℃ 2 h, and the polyclonal antibody working time was 50 min. The standard curve was established under the optimal conditions, the minimum detectable limit was 0.1 μg?mL ^-1. This method was specific, with no cross reaction with HBHA, FO, FV, and AO. Meanwhile, the inter-assay repetition rate was 1.5%-5.8% and intra-assay repetition rate was 0.4%-3.2%, both lower than 10%, indicating it was good repeatability. Non-aflatoxigenic fungi had lower values, close to negative value. 【Conclusion】 The large-capacity reaction system sandwich ELISA method established in this study could quickly and accurately detect aflatoxigenic fungi, which laid a foundation for further control of aflatoxin contamination from the source.

Sequence Structure and Expression Characteristics Analysis of AGTR2 in Bovine Follicle

ZhiWei ZHU,ShuNing HOU,QingLing HAO,JiongJie JING,LiHua LÜ,PengFei LI

Scientia Agricultura Sinica. 2020, 53(7): 1482-1490. doi:10.3864/j.issn.0578-1752.2020.07.016

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【Objective】 The aim of study was to identify molecular characteristics and three-dimensional structure of AGTR2 in the bovine follicular development, and the function was analyzed by combining expression characteristics of AGTR2 in different physiological follicles.【Method】Ovaries in the bovine follicular were observed by B-type ultrasonography (twice a day), removed from cows when DF and SF appeared, and then separated DF and SF; GCs were isolated, total RNA was extracted and detected by RT-PCR, specific primers were amplified and sequenced, and CDS region sequence structure was obtained; the bioinformatics method was used to analyze its sequence structure, relationship and three-dimensional structure; the qRT-PCR primers of AGTR2 and reference gene RPLP0 were designed to analyze differential expression level of AGTR2 in DF and SF; DF and SF of another cow were fixed with 4% paraformaldehyde, positive, negative control groups and experimental groups were set, expression level and localization of AGTR2 were analyzed by immunohistochemistry. 【Result】The results showed that total length of AGTR2 CDS region was 1 089 bp, encoding 362 amino acids; BLAST analysis of amino acid sequence of AGTR2 and the corresponding amino acid sequence of other 24 animals obtained by NCBI database indicated that the sequence had the highest similarity with buffalo (99.4%) and 92.0%-98.9% with other animals; The three-dimensional structure and functional domain analysis showed that AGTR2 possessed 7 parallel alpha helical structures across the cell membrane, which was a typical G protein-coupled receptor; the results of qRT-PCR analysis showed that expression level of AGTR2 mRNA in DF was significantly higher than SF (P<0.01), and the differential expression multiple was up to 7.47 times. The immunohistochemical analysis showed that AGTR2 was expressed in GCs and membrane cells layer of DF and SF, and the specific color intensity showed that expression of AGTR2 in SF membrane cells was higher than DF. 【Conclusion】AGTR2 belonged to G protein-coupled receptor and conforms to basic characteristics of CART receptor, and the study laid a foundation for further study on mechanism of AGTR2 regulating signal pathway and hormone secretion during bovine follicular development, meanwhile, it was of great significance for identification of CART receptor and in-depth explanation of mechanism of CART regulating bovine follicular development.

Genome-Wide Identification and Characterization of Transposable Elements in Xichuan Black-Bone Chicken

DongHua LI,YaWei FU,ChenXi ZHANG,YanFang CAO,WenTing LI,ZhuanJian LI,XiangTao KANG,GuiRong SUN

Scientia Agricultura Sinica. 2020, 53(7): 1491-1500. doi:10.3864/j.issn.0578-1752.2020.07.017

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【Objective】The aim of the study was to analyze the whole genome re-sequencing data of Xichuan black-bone chicken to obtain the identification, classification, distribution of the transposon of Xichuan black-bone chicken, and to explore the pathways involved in transposon-related genes, which not only had important significance for studying the biological function of the Xichuan black-bone chicken transposon elements (TEs), but also provided important basic data for exploring genome amplification, genome function and evolution research. 【Method】In this study, whole genome resequencing of blood DNA of Xichuan black-bone chicken was performed, and the paired-end mapping methods alignment genome was used. The TEs was identified, constructed, corrected and classified by RepeaterModeler, TEclass, RepeatMasker and other processes. All TEs in the whole genome of Xichuan black-bone chicken were obtained to analyze the characteristics, distribution and relationship of their genes. All genes inserted in the TEs were subjected to GO and KEGG databases enrichment, and the functions were described in combination with GO and KEGG annotation results, respectively. 【Result】 After identification, classification and annotation, 370 252 TEs sequences were identified in Xichuan black-bone chicken, which were divided into 19 superfamilies, mainly CR1, TcMar-Mariner, ERVL, ERV1 and other superfamilies, further indicated that the TEs types of Xichuan black-bone chicken were major TEs. The number of TEs was related to chromosome length, and longer the chromosome was, the more the number of TEs was. Number of TEs was inversely proportional to the density of gene. TEs density was relatively low in gene dense regions. The insertion of TEs in genome was not random, LTR/ERVL, LTR/ERV1, DNA/PIF-Harbinger, DNA/Hat-Charlie, RC/Helitron tend to be inserted outside the gene. GO enrichment analysis indicated some of these genes related to TEs were enriched in the following biological process terms: cell process, single-organism process, metabolic process, biological regulation, response to stimulus. In addition, some of these genes related to TEs were enriched in the following molecular function terms: binding and catalytic activity. Furthermore, some of these genes related to TEs were enriched in the following cell component terms: cell parts, organelles, and membranes. KEGG enrichment analysis indicated that these genes related to TEs mainly focused on Glycerolipid metabolism, PPAR signaling pathway, PI3K-Akt signaling pathway, Insulin resistance, and Jak-STAT signaling pathway. This study mainly focused on the pathways related to the characteristics of Xichuan black-bone chicken, such as tyrosine metabolism related to pigmentation, lipid metabolism related to meat quality, fatty acid synthesis, and PI3K-Akt signaling pathway. 【Conclusion】 There was a positive correlation between TEs content and genome size of Xichuan black-bone chicken. Moreover, the TEs of Xichuan black-bone chicken had a certain preference in the distribution of genome. TEs related genes were enriched in the pigmentation related pathway, which might be related to the germplasm characteristics of Xichuan black-bone chicken. The specific regulatory mechanisms remained to be further studied.

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