Table of Content

    01 March 2023, Volume 56 Issue 5
    Scientia Agricultura Sinica. 2023, 56(5):  0. 
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    Genome-Wide Association Studies and Mining for Favorable Loci of Root Traits at Seedling Stage in Wheat
    WANG Mai, DONG QingFeng, GAO ShenAo, LIU DeZheng, LU Shan, QIAO PengFang, CHEN Liang, HU YinGang
    Scientia Agricultura Sinica. 2023, 56(5):  801-820.  doi:10.3864/j.issn.0578-1752.2023.05.001
    Abstract ( 213 )   HTML ( 56 )   PDF (7962KB) ( 195 )   Save
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    【Objective】Plant roots are critical for water and nutrient acquisition, crop growth and development as well as yield formation. Exploring SNP loci significantly associated with root traits in wheat at seedling stage and mining candidate genes, will lay a foundation for understanding the genetic mechanism of wheat root system architecture and breeding wheat elite varieties with better root architecture.【Method】In this study, 189 diverse wheat cultivars were assembled as an association-mapping panel, five root traits including total root length (TRL), total root area (TRA), total root volume (TRV), average root diameter (ARD) and root dry weight (RDW) were investigated by growing in two culture conditions (Hoagland nutrient solution and pure water), and the experiments were repeated twice. Then, genome-wide association studies (GWAS) were performed for the five root traits with genotypic data derived from Wheat 660K SNP Array. Candidate genes were predicted by sequence alignment, domain analysis, and annotation information. Futhermore, kompetitive allele specific PCR (KASP) markers were developed for root traits. 【Result】The root traits varied greatly among the 189 cultivars, and the roots were thick and short cultured under Hoagland nutrient solution, while slender seminal roots and more lateral roots were observed under pure water. A total of 95 QTLs significantly associated with root traits cultured in two conditions (P<10-3) were identified by genome-wide association studies with four models of BLINK (bayesian-information and linkage-disequilibrium iteratively nested keyway), CMLM (compressed mixed linear model), FarmCPU (fixed and random model circulating probability unification) and MLM (mixed linear model). Among them, 18 QTLs were detected in both culture conditions and distributed on chromosomes of 7A, 1B, 2B, 3B, 7B, 1D, 2D, and 3D, which explained 8.68%-14.07% of phenotypic variation. Of those significant loci, 4 QTLs were similar or consistent with that reported previously, and the rest were novel ones. Haplotype analysis conducted for co-localization QTLs of 10 SNPs revealed significant differences in root traits between the two haplotypes of wheat cultivars. Based on these SNPs, KASP markers XNR7143 and XNR3707 were developed for total root volume and root dry weight, respectively. In addition, 12 candidate genes possibly regulating root development were found by mining the genes within the interval of co-localization significant SNPs. Of them, TraesCS7A02G160600, encoding 3-oxoacyl-[acyl-carrier-protein] synthase, is involved in the synthesis of root fatty acids; TraesCS1B02G401800, encoding syntaxin, plays an important role in plant tropism; TraesCS7B02G417900, encoding aldehyde oxidase, contributes to the synthesis of abscisic acid and regulation of crop root development. 【Conclusion】The root traits of wheat varied significantly among the wheat genotypes. Genome-wide association studies detected 18 significant QTLs linked with root traits simultaneously in two culture conditions, two KASP markers were developed for root traits, and 12 candidate genes related to root development were screened, which might provide reference for understanding the regulation mechanism of wheat root traits and molecular marker-assisted breeding for wheat improvement.

    Candidate Gene Localization of ZmDLE1 Gene Regulating Plant Height and Ear Height in Maize
    ZHOU WenQi, ZHANG HeTong, HE HaiJun, GONG DianMing, YANG YanZhong, LIU ZhongXiang, LI YongSheng, WANG XiaoJuan, LIAN XiaoRong, ZHOU YuQian, QIU FaZhan
    Scientia Agricultura Sinica. 2023, 56(5):  821-837.  doi:10.3864/j.issn.0578-1752.2023.05.002
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    【Objective】 Plant height is one of the important target traits in maize plant type breeding, which is closely related not only to mechanized grain harvesting and lodging resistance, but also to maize yield. Therefore, it is of great theoretical and breeding value to isolate QTL/gene of maize plant height and analyze its function. This study aims to locate a novel maize dwarf gene ZmDLE1, clarify its biological function, and provide important theoretical basis and gene resources for accelerating the improvement of maize plant type. 【Method】 A single recessive mutant was derived in maize inbred line LY8405, from Crop Research Institute of Gansu Academy of Agricultural Sciences, by chemical mutagenic agent Ethyl Methyl Sulfonate (EMS). A maize dwarf and low ear mutant was isolated from the M2 progeny, and the M3 and M4 progeny could stably inherit, which was named dwarf and low ear mutant1 (Zmdle1). The F2 population was constructed by hybridization with Mo17 and was identified by bulked segregate analysis-sequencing (BSA-seq) and target segment recombination exchange. Based on the Mo17 reference genome, the genes in the target region were obtained and functionally annotated to locate candidate genes.【Result】Phenotypic identification of Zmdle1 was carried out, and the phenotype of Zmdle1 at seedling stage was not significantly different from that of the control LY8405. The plant height and ear height of Zmdle1 at mature stage were reduced by 87.2 cm and 55.4 cm, respectively, accounting for about 35.0% and 62.9%, difference is extremely significant. Morphological observation showed that the decrease of internode number and the shortening of internode cell length were the main reasons for the significant decrease of plant height and ear height of Zmdle1. The genetic analysis of the mutant gene was conducted by using the F2:3 genetic populations. The Zmdle1 mutant is inherited in a 3﹕1 (χ2=2.854) ratio and is a single recessive gene. Therefore, according to the results of BSA-seq, the candidate gene ZmDLE1 was initially located in the 15 Mb region of Bin1.09-1.10 on chromosome 1 of maize. The polymorphism molecular markers were further developed using the re-sequencing results of Mo17 and Zmdle1, and the target gene was accurately cloned by map-based cloning. Finally, the candidate genes were mapped to the size range of 600 kb, and there were 16 candidate genes in this range. By comparing the re-sequencing data, it was found that Zm00001d033231 gene changed into A at the 2062 position G, which resulted in amino acid changing from glycine to serine, and the transcription level expression was significantly reduced compared with LY8405. Zm00001d033234 changed from T to C at the 223rd position leading to the 75th amino acid changed from serine to proline, and there was no significant difference in transcription level. Through association analysis of natural populations and the predicted genes for functional annotation, it was found that Zm00001d033231 and Zm00001d033234 were related to the growth and development of maize. 【Conclusion】 The candidate gene ZmDLE1 in the Bin1.09 region at the end of chromosome 1, was identified to effectively regulate maize plant height and ear height, and the target region was reduced to 600 kb by fine localization. Association analysis showed that Zm00001d033231 was genetic locus significantly associated with plant height within the target region.

    Response of Water Use Characteristics of Spring Wheat to Co- Incorporation of Green Manure and Wheat Straw in Arid Irrigation Region
    FAN ZhiLong, HU FaLong, YIN Wen, FAN Hong, ZHAO Cai, YU AiZhong, CHAI Qiang
    Scientia Agricultura Sinica. 2023, 56(5):  838-849.  doi:10.3864/j.issn.0578-1752.2023.05.003
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    【Objective】The aim of this study was to investigate the effects of green manure and wheat straw co-incorporation on water use efficiency of next season wheat (Triticum aestivum L.), which played an critical supporting role in the construction of water use efficient and sustainable production pattern of spring wheat in northwest irrigation region.【Method】In this study, a randomized block design was adopted, with fallowing after wheat harvesting (W) as the control, wheat multiple cropping green manure with no-tillage sowing + green manure and wheat straw co-incorporation (W-NTGS), wheat multiple cropping green manure with no-tillage sowing + green manure incorporation (W-NTG), and wheat multiple cropping green manure shallow plowing for destroying stubble + single returning of green manure to the field (W-TG). The characteristics of water consumption, grain yield, water use efficiency (WUE) and irrigation water use efficiency (WUEi) of wheat were investigated. 【Result】 The soil water storage in 0-50 cm soil layer before wheat sowing was significantly increased by multiple cropping green manure compared under W. The soil water storage in 0-10 cm, 10-20 cm, 20-30 cm, and 30-50 cm soil layers before wheat sowing with W-NTGS was increased by 22.1%, 30.2%, 21.5%, and 11.1% compared with that of W-NTG, and 26.2%, 33.2%, 26.5%, and 16.4% increase compared with that of W-TG, respectively. However, there was no significant difference in soil water storage in 0-110 cm soil layer before wheat sowing between W-NTG and W-TG. Compared with fallow treatment, water consumption, evaporation, and ratio of evaporation to evapotranspiration of wheat decreased by 7.0%-7.1%, 11.7%-20.1%, and 5.2%-15.9%, respectively. The water consumption of wheat with W-NTGS decreased by 6.4% than that of wheat under W-NTG in 2021, and averagely decreased by 6.1% than that of wheat under W-TG in 2020 and 2021. Compared with W-NTG and W-TG, the evaporation of wheat under W-NTGS decreased by 9.7% and 13.6% on average, and the ratio of evaporation to evapotranspiration decreased by 6.2% and 11.3%, respectively. There were no significant differences in water consumption, evaporation, and evapotranspiration ratio between W-NTG and W-TG. Compared with W, the grain yield of wheat multiple cropped with green manure increased by 6.4%-16.8%. Meanwhile, the grain yield of wheat under W-NTGS increased by 6.6% and 9.8% on average compared with W-NTG and W-TG, respectively. However, the grain yield of wheat between with W-NTG and W-TG was not significant difference. Compared with W, the WUE and WUEi of wheat multiple cropped with green manure increased by 11.9%-30.7% and 6.4%-16.8%, respectively. The WUE of wheat under W-NTGS increased by 10.9% and 16.8%, and the WUEi increased by 6.6% and 9.8%, respectively compared with W-NTG and W-TG. There were no significant differences in WUE and WUEi between W-NTG and W-TG.【Conclusion】 The co-incorporation of green manure and wheat straw (W-NTGS) could significantly increase soil water storage in 0-50 cm before wheat sowing, and decrease the ineffective loss of soil water in wheat season, thus reducing the water consumption of wheat and increasing the grain yield, and ultimately significantly improving the farmland and irrigation water use efficiency, which could be used as a recommended technology for water efficient use of wheat multiple cropping with green manure in arid irrigation region.

    UAV Multispectral Image-Based Nitrogen Content Prediction and the Transferability Analysis of the Models in Winter Wheat Plant
    GUO Yan, JING YuHang, WANG LaiGang, HUANG JingYi, HE Jia, FENG Wei, ZHENG GuoQing
    Scientia Agricultura Sinica. 2023, 56(5):  850-865.  doi:10.3864/j.issn.0578-1752.2023.05.004
    Abstract ( 138 )   HTML ( 21 )   PDF (1290KB) ( 116 )   Save
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    【Objective】Accurate monitoring and rational application of nitrogen are particularly important for healthy growth, yield and quality improvement of wheat, and reduction of environmental pollution and resource waste. The purpose of this study was to develop UAV-based models for accurately and effectively assessment of the plant nitrogen content in the key growth stages of wheat growth, and to explore the transferability of the models constructed based on machine learning methods. 【Method】Winter wheat experiment were conducted from 2020 to 2022 in Shangshui county, Henan province, China. Based on the K6 multichannel imager mounted on DJM600 UAV, 5-band (Red, Green, Blue, Rededge, and Nir) multispectral images were obtained from a UAV system in the stages of jointing, booting, flowering and filling in winter wheat, to calculate 20 vegetation indices and 40 texture features from different band combinations. Correlation analysis was used to screen the sensitive characteristics of nitrogen content in winter wheat plants from the 65 image features. Combining the sensitive spectral features and texture features of the nitrogen content of winter wheat plants, BP neural network (BP), random forest (RF), Adaboost, and support vector machine (SVR) machine learning regression methods were used to build plant nitrogen content models, and compared for the model performance and transferability. 【Result】(1)The correlation coefficients between plant nitrogen content and image features passed the test of 0.01 extremely significant level, including 22 spectral features and 29 texture features. (2) 51 spectral and texture features were adopted to build four machine learning models. The estimates of plant nitrogen by the RF and Adaboost methods were relatively concentrated, mostly close to the 1﹕1 line; while the estimations from the BP and SVR methods were relatively scattered. The RF method was the best, with R2, RMSE, and MAE of 0.81, 0.42%, and 0.29%, respectively; The SVR method was the worst, with R2, RMSE, and MAE of 0.66, 0.54% and 0.40%, respectively. (3) The prediction effects of the four methods on the nitrogen content of W0 and W1 treatments trained using W1 and W0 treatments were the same as those trained using both W0 and W1 datasets, both of which were closer to the 1﹕1 line for the RF and Adaboost methods. The R2 of transfer prediction results for the models constructed by BP, RF, Adaboost, and SVR methods were 0.75, 0.72, 0.72, and 0.66 for the prediction of nitrogen content in W0 treatment and 0.51, 0.69, 0.61 (trained using data under W1 treatment) and 0.45 for the prediction under W1 treatment (trained using data under W0 treatment), respectively.【Conclusion】All models showed strong transferability, especially the RF and Adaboost methods, in predicting winter wheat nitrogen content under rainfed and irrigation water management.

    Functional Analysis of Effector Hasp83 in the Pathogenicity of Puccinia striiformis f. sp. tritici
    WANG JianFeng, CHENG JiaXin, SHU WeiXue, ZHANG YanRu, WANG XiaoJie, KANG ZhenSheng, TANG ChunLei
    Scientia Agricultura Sinica. 2023, 56(5):  866-878.  doi:10.3864/j.issn.0578-1752.2023.05.005
    Abstract ( 133 )   HTML ( 21 )   PDF (1980KB) ( 78 )   Save
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    【Background】Wheat stripe rust is a serious disease on wheat, which is caused by Puccinia striiformis f. sp. tritici (Pst). Pst is an obligate biotrophic fungus, which can form haustorium during infection and absorb nutrients from the host plants via haustorium. Moreover, Pst secretes effectors through haustorium to regulate host immunity and promotes the infection process.【Objective】The objective of this study is to clarify the function and mechanism of Pst effectors, and to reveal the pathogenicity mechanism of Pst.【Method】By comparing the transcriptome of Pst urediospore, germinated tube and haustorium, Hasp83 encoding secreted protein was identified to be significantly induced in haustorium, and whether it could inhibit the cell death caused by BAX on Nicotiana benthamiana leaves was observed through Agrobacterium-mediated transient expression. qRT-PCR was used to detect the expression level of Hasp83 during different Pst infection stages in wheat. The type Ⅲ secretion system (T3SS) of Pseudomonas fluorescens EtHan and host induced gene silencing (HIGS) were carried out to investigate the function of Hasp83 during Pst infection. The yeast two-hybrid (Y2H) system was used to screen the proteins interacting with Hasp83 in wheat, and co-inmunoprecipitation (Co-IP) assay was used to further verify the interaction by co-expressing Hasp83 and its candidate target proteins in N. benthamiana cells.【Result】The open reading frame (ORF) of Hasp83 is 522 bp, encoding 173 amino acids. Hasp83 contains no conserved domain, and the N-terminal 1-29 amino acids encode a signal peptide, which could inhibit the cell death caused by BAX on N. benthamiana leaves through Agrobacterium-mediated transient expression. qRT-PCR analysis revealed that Hasp83 was up-regulated during Pst infection in wheat. Transient expression of Hasp83 in wheat via T3SS could inhibit callose accumulation caused by P. fluorescens, and lead to 19.35%-38.62% decrease of reactive oxygen species (ROS) accumulation area and necrotic cell area caused by the avirulent Pst race CYR23 in wheat. Silencing of Hasp83 by HIGS in Suwon11 wheat leaves infected with the virulent Pst race CYR31 significantly reduced pathogenicity of Pst compared to controls, resulting in less urediospore sporulation, shorter infection hyphal length, smaller infection area, and decreased haustorium number. Y2H result showed that the effector Hasp83 interacted with wheat hypersensitive-induced reaction (HIR) protein, Tahir1. The interaction between Hasp83 and Tahir1 was further confirmed by Co-IP assay in N. benthamiana.【Conclusion】Pst effector Hasp83 can suppress wheat immunity caused by the non-pathogenic bacteria and avirulent Pst, and enhance the pathogenicity of Pst.

    Expression and Functional Analysis of Glycosyl Hydrolase Genes from Plasmopara viticola
    PAN FengYing, QU JunJie, LIU LuLu, SUN DaYun, GUO ZeXi, WEI XiaoLi, WEI ShuMei, YIN Ling
    Scientia Agricultura Sinica. 2023, 56(5):  879-891.  doi:10.3864/j.issn.0578-1752.2023.05.006
    Abstract ( 109 )   HTML ( 24 )   PDF (3373KB) ( 82 )   Save
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    【Objective】The objective of this study is to clone glycosyl hydrolase genes from Plasmopara viticola (PvGHs), analyze their characteristics and expression patterns in infected grape leaves, and the abilities to inhibit or promote programmed cell death (PCD) and affect Phytophthora nicotianae infection in tobacco leaves, so as to provide a theoretical basis for further study on mechanism of regulating host plant immunity.【Method】Eight PvGHs with full-length were amplified by RT-PCR from P. viticola Pv5-27 strain. The sequences of these eight PvGHs and encoded proteins were analyzed by bioinformatics. Yeast signal peptide trap system (SST) was used to verify the secretory activity of the PvGH proteins. The expression pattern of PvGHs during infection grape leaves was detected by qRT-PCR. At the same time, eight PvGH effector proteins were transiently expressed in Nicotiana benthamiana by Agrobacterium-mediated PVX virus expression system. Moreover, their inhibitory abilities to inhibit INF1- and BAX-triggered PCD and to promote P. nicotianae infection were also analyzed.【Result】The sequences of eight PvGHs were completely consistent with the prediction in genome sequence, with the length of 1 092 to 1 392 bp, encoding 364-464 amino acids, respectively. The similarity with homologous proteins from other oomycetes was as high as 60.62%-86.36%. None of them had transmembrane domains. Their secondary structures were quite different from each other, and their tertiary structures were less similar to those of other proteins, which showed a very unique tertiary structure. SingalP 5.0 software was used to predict signal peptide of these proteins. It was found that all the PvGH proteins contained signal peptide sequences of 20-26 aa in length. However, the SST verification results showed that PvG09279 and PvG13517 do not have secretion activity. The retention, deletion or replacement of signal peptide sequences of the eight PvGH proteins could inhibit the BAX-triggered PCD and all these PvGH proteins could promote P. nicotianae infection in tobacco leaves. It suggests that the potential virulence of eight PvGH effectors does not dependent on the signal peptide. The up-regulated expression of PvGHs in the early stage of infection of P. viticola further indicated that PvGHs play an important role in the interaction between pathogen and host.【Conclusion】During downy mildew infection, PvGHs secreted by P. viticola are involved in pathogenic process by inhibiting the host PTI response.

    Effect of Subsoiling Combined with Straw Returning Measure on Pore Structure of Black Soil
    YANG JianJun, GAI Hao, ZHANG MengXuan, CAI YuRong, WANG LiYan, WANG LiGang
    Scientia Agricultura Sinica. 2023, 56(5):  892-906.  doi:10.3864/j.issn.0578-1752.2023.05.007
    Abstract ( 116 )   HTML ( 19 )   PDF (2747KB) ( 82 )   Save
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    【Objective】The effect of subsoiling combined with straw returning on soil pore structure of black soil in Northeast China is lack of definite judgment. Aimed at such problem, this research was conducted in order to provide scientific basis for the influence mechanism of this measure on soil structure of black soil and the establishment of reasonable tillage.【Method】In this study, a 5-year field positioning experiment conducted in Qinggang County, Suihua City, Heilongjiang Province, a typical black soil area in Northeast China was used as a platform. Farmers’ conventional treatment (FP), 25 cm subsoiling alone treatment (T2), 25 cm subsoiling combined with straw returning treatment (T3) and 35 cm subsoiling combined with straw returning treatment (T4) were set. The visualization and quantification of soil pore structure were studied using CT scanning technology, and combined with field capacity and bulk density to explore the effect of subsoiling combined with straw returning on the pore structure of black soil. 【Result】The results showed that the two-dimensional and three-dimensional images of soil pores clearly showed that the pore distribution at 0-20 cm soil layer was significantly less than that at 20-40 cm soil layer in all treatments, while the pore distribution of suboiling combined with straw returning treatment (T3 and T4) was significantly higher than that under FP treatment, and their macropores with more complex structure were increased. Quantitative analysis showed that compared with FP treatment, the total porosity of 20-30 cm soil layer under T2 was significantly increased by 103.0% (P<0.05), which was achieved by significantly increasing the micropores porosity(pore diameter d≤0.50 mm) by 91.3% and mesopores porosity (0.50 mm<d≤1.00 mm) by 143.5% (P<0.05). While subsowing combined with straw returning treatments (T3 and T4) significantly increased the total porosity of 0-30 cm soil layer by 109.8%-382.7% (P<0.05), which was achieved by significantly increasing the macropores porosity (d>1.00 mm) by 221.5%-661.7% and the mesopores porosity by 105.4%-544.9% (P<0.05). In addition, compared with FP treatment, subsoiling combined with straw returning (T3 and T4) significantly increased the fractal dimension of soil pores at 0-30cm soil layer by 9.9%-17.7% (P<0.05), decreased the Euler number by 32.4%-66.4% (P<0.05), and significantly increased the field water capacity by 24.2%-40.6% (P<0.05). Further analysis showed that the different pore size porosity and total porosity was significantly positively correlated with field capacity and fractal dimension, but significantly negatively correlated with Euler number (P<0.01). 【Conclusion】Subsoiling combined with straw returning could improve porosity of macropores and mesopores pore of black soil, improve pore structure and connectivity, and increase field capacity, especially the effect of 35 cm subsoiling combined with straw returning treatment is the most significant, which could be recommended measure for rational tillage construction in black soil of Northeast China.

    Effects of Long-Term Conservation Tillage on Soil Carbon Content and Invertase Activity in Dry Farmland on the Loess Plateau
    Scientia Agricultura Sinica. 2023, 56(5):  907-919.  doi:10.3864/j.issn.0578-1752.2023.05.008
    Abstract ( 127 )   HTML ( 20 )   PDF (1118KB) ( 70 )   Save
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    【Objective】The effects of long-term conservation tillage on soil carbon and carbon invertase activity in forage-crop rotation system of dry farmland in the Loess Plateau were explored to provide the scientific basis for soil carbon sequestration and sustainable and healthy development of agriculture in dry farmland. 【Method】In this study, we aimed to investigate the effects of long-term traditional tillage (T), no-tillage (NT), traditional tillage+straw mulch (TS), and no-tillage+straw mulch (NTS) on soil organic carbon (SOC), microbial biomass carbon content (MBC), β-glycosidase (βG), cellobiohydrolase (CBH) and β-xylosidase enzymes (βX) in forage-crop rotation system at the National Field Scientific Observation and Research Station of the Grassland Agricultural Ecosystem in Qingyang, Gansu Province. Soils were collected from 0-5, 5-10 and 10-20 cm depths at the harvest of maize (Zea mays L.). 【Result】(1) Conservation tillage significantly increased the contents of SOC and MBC in soil, especially in 0-5 cm soil layer. Compared with conventional tillage, straw mulching increased SOC and MBC by 19.1% and 39.9%, respectively, and no-tillage increased SOC and MBC by 15.1% and 34.3%, respectively. (2) Conservation tillage significantly increased soil carbon invertase activity, the three enzyme activities showed: βG>CBH>βX, the sensitivity of conservation tillage measures showed: CBH>βX>βG. Compared with traditional tillage, the activities of βG, CBH and βX in 0-5 cm soil layer under straw mulching increased by 20.3%, 37.6% and 41.1%, respectively, and those under no-tillage increased by 12.5%, 31.0% and 26.1%, respectively. Straw mulching in 5-20 cm soil layer increased βG, CBH and βX by -7.6%, 99.9% and 3.5%, respectively, and no-tillage increased them by -21.1%, 22.1% and -12.1%, respectively. In addition, the structural equation results showed that soil carbon invertase activity was mainly affected by soil total nitrogen content in forage-crop rotation system. (3) Straw mulching could directly affect the accumulation of soil carbon, mainly affecting the activity of soil carbon invertase by changing the soil total nitrogen content; no tillage had no significant impact on soil environment, resulting in no significant changes in soil carbon content and carbon invertase activity.【Conclusion】The accumulation of soil carbon was mainly affected by the direct effect of straw mulching measures, and the activity of carbon invertase was mainly changed by the indirect effect of soil total nitrogen in the forage-crop rotation system of dry farmland in the Loess Plateau. Among them, no tillage combined with straw mulching was the most effective measure to improve soil carbon content and enzyme activity, and β- Glucosidase is the main enzyme involved in soil carbon inversion.

    Environmental Risks Assessment of Livestock and Poultry Non- Point Source Pollution in Shiyan City Based on Arable Land Carrying Capacity
    GONG ShiFei, XIAO NengWu, DING WuHan, JU XueHai, WU PingHua, YU YongSong, LI Hu
    Scientia Agricultura Sinica. 2023, 56(5):  920-934.  doi:10.3864/j.issn.0578-1752.2023.05.009
    Abstract ( 109 )   HTML ( 42 )   PDF (578KB) ( 47 )   Save
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    【Objective】 Shiyan city was the core water source area of the middle route of the South-to-North Water Transfer Project, a comprehensive understanding of the current situation of livestock and poultry pollution and its potential threat to farmland would provide a basis for strengthening environmental risk control. 【Method】This paper used the livestock manure environmental risk assessment model to quantitatively analyze the source structure of livestock and poultry manure in different areas of Shiyan city in 2020, and the environmental risk from the perspective of the balance between planting and raising was comprehensively evaluated. 【Result】 The total amount of livestock and poultry manure reached 306.85×104 t in 2020, and it was mainly distributed in Yunyang district, Danjiangkou city and Fangxian county. COD production was the highest (57.00×104 t), followed by TN (2.63×104 t), and TP production was less (0.65×104 t). The total equivalent standard pollution load was 15.532×1010 m3, and the source structure was obviously inconsistent in different regions. TP was the primary pollutant in livestock and poultry breeding, and its equivalent standard pollution load accounts for 41.68% of the total load, followed by TN with the pollution load rate of 33.85%, and the lowest equivalent standard pollution load rate of COD was 24.47%. Cattle was the primary source of non-point source pollution in livestock and poultry production in Shiyan city, contributing 49.47% to the total pollution load of the equivalent standard, followed by poultry (21.60%). According to the result of environmental pollution risk assessment, the load of animal excretion per hectare of arable land in Shiyan city was 10.15 t·hm-2, the alarm value was 0.338 and the risk level was Ⅰ, indicating the environmental pollution was “no risk”. The load intensity of TN and TP was 0.09 and 0.02 t·hm-2, respectively, and the environmental risk index was 1.023 and 1.223, respectively, but there were obvious spatial differences among regions. More than half of the whole city had the potential pollution risk of livestock and poultry breeding, among which Maojian district and Yunyang District were the most serious. With phosphorus as the measurement standard, the total amount of breeding in Shiyan city was close to saturation, the total amount of livestock and poultry must be reduced by 98.51×104 pigs equivalent on the existing basis.【Conclusion】It was of great significance to maintain a reasonable scale of poultry breeding (≤441.00×104 pigs equivalent) and take measures of pollutant abatement, in order to promote the balance of planting and breeding and orderly development of livestock and poultry breeding in Shiyan city.

    Development and Validation of KASP Markers Based on a Whole- Genome Resequencing Approach in a Hybrid Population of Luli × Red No. 1
    ZHENG WenYan, CHANG YuanSheng, HE Ping, HE XiaoWen, WANG Sen, GAO WenSheng, LI LinGuang, WANG HaiBo
    Scientia Agricultura Sinica. 2023, 56(5):  935-950.  doi:10.3864/j.issn.0578-1752.2023.05.010
    Abstract ( 100 )   HTML ( 20 )   PDF (2808KB) ( 64 )   Save
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    【Objective】 A series of kompetitive allele-specific PCR (KASP) markers for traits related to apple leaves were developed to provide a reference to effectively utilize light in apple breeding using a Luli ×Red No. 1 hybrid population as materials. 【Method】This study investigated the leaf trait phenotypic data (including leaf length, width, contents of chlorophyll and anthocyanin, photosynthetic rate and chlorophyll fluorescence parameters) of Luli, a type I red-fleshed apple, Red No. 1, and 134 hybrids. Whole-genome resequencing was conducted on an Illumina HiSeq 2500 platform. The clean sequencing reads were mapped to the apple reference genome using BWA software. GATK software was used to identify single nucleotide polymorphisms (SNPs). 【Result】A total of 164 776 660, 149 482 876, and 3 927 370 200 clean reads were obtained by whole-genome resequencing from Luli, Red No. 1 and 134 hybrid individuals, respectively. The data were aligned to the reference genome sequences with mapping rates of 98.77%, 98.89%, and 97.79%, respectively. A total of 6 445 766 SNPs were identified. The SNPs were then filtered, and 94 208 accurate and high-quality SNPs were selected for subsequent KASP primer design. Finally, 5 802 KASP markers were developed based on a high-throughput whole-genome resequencing approach. The average polymorphism information content (PIC) of these KASP markers was 0.31, and there were 30.13% PICs>0.35. The Simpson genetic diversity index ranged from 0.01 to 0.67, with an average value of 0.53. The mean observed heterozygosity was 0.32. A Kyoto Encyclopedia of Genes and Genomes enrichment analysis of the genes for KASP marker revealed that “one carbon pool by folate” was the most significant pathway. An analysis of the phenotypic data of leaf-related traits in both the parents and individuals from the Luli ×Red No. 1 cross showed that there were significant differences in the phenotypic values of leaf net photosynthetic rate, stomatal conductance (Gs) and other traits between the parents Luli and Red No. 1. The F1 generation exhibited a wide phenotypic variation, and the most traits segregated extensively. Α detailed study of the phenotypic data of leaf traits of the hybrid population with the genotype and annotations of the KASP markers finally resulted in the development of 21 KASP markers, which was correlated with leaf-related traits, including chlorophyll content, Gs and leaf length and width.【Conclusion】Use of the whole-genome resequencing data of Luli, Red No. 1 and 134 hybrid individuals resulted in the development of 5 802 KASP. A total of 21 of these KASP markers significantly correlated with leaf traits, which provided a theoretical reference for the high utilization of light energy in apple breeding.

    Genomic Selection for Fruit Weight and Soluble Solid Contents in Peach
    CAO Ke, CHEN ChangWen, YANG XuanWen, BIE HangLing, WANG LiRong
    Scientia Agricultura Sinica. 2023, 56(5):  951-963.  doi:10.3864/j.issn.0578-1752.2023.05.011
    Abstract ( 110 )   HTML ( 17 )   PDF (1706KB) ( 47 )   Save
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    【Background】 Fruit weight and soluble solid content (SSC) are two important quantitative traits in peach which are of importance to breeders. However, performing early prediction using a single marker is challenging as the traits are controlled by multiple minor genes. Genomic selection, a novel genome-wide tool, has been applied in fruit crops and can potentially enhance the breeding efficiency of these quantitative traits. However, its effects in peach and influencing factors require further investigation.【Objective】 Establish a whole-genome selection technology system for peach single fruit weight and SSC, and laid a methodological foundation for the establishment of efficient molecular breeding technology system for peach.【Method】 The objectives of this study were to assess the accuracy of prediction of peach fruit weight and SSC in natural and hybrid populations using genomic selection. Here, a training population of 520 individuals was selected. Using genotypic data for 48 398 single nucleotide polymorphisms (SNPs) obtained from the resequencing results of the above training population, a total of 11 genome-wide prediction models were built to select the optimum model for fruit weight and SSC. Subsequently, the genomic breeding values of a small natural population of 56 individuals and 29 hybrid populations comprising a total of 1 145 seedlings were calculated.【Result】 The average sequencing data of each variety of the three groups was 1.95-3.52 Gb, and the sequencing depth was 5.29-10.79×. The sequencing data of the training natural population was aligned with the reference genome, and a total of 5 065 726 SNPs were obtained. After removing the SNPs with a high missing rate (>20%) and minor allele frequency of <0.05, a total of 48 398 SNPs on the genome were randomly selected for constructing whole-genome selection models for the training population. The models with the highest prediction accuracy for fruit weight and SSC were BayesA and randomforest, respectively. Using the above two models, it was found that the goodness of fit between the predicted breeding values and observed phenotype of fruit weight was 0.4767-0.6141, which was higher than that of SSC (0.3220-0.4329) in the natural populations. In hybrid populations, the prediction accuracy of fruit weight was 0.2319-0.4870, which was also higher than that of SSC (0.0200-0.2793). The results also showed that the prediction model constructed by training natural populations was more accurate in predicting natural populations than hybrid populations. Taking fruit weight as an example, it was also found that only 17.78% of the seedlings needed to be retained by genomic selection when targeting large fruit. Genomic selection was significantly more efficient than single and double marker selection. Furthermore, the effects of population dispersion, heritability and population structure on prediction accuracy are also discussed. The results indicated that prediction accuracy may vary and be affected by a combination of several factors.【Conclusion】 In this study, a suitable genomic selection model for peach fruit weight and SSC was screened, and it was confirmed that the prediction efficiency of genomic selection was significantly higher than that of single marker selection. The results indicated the potential of genomic prediction in accelerating breeding progress of these two quantitative traits in peach.

    Transcriptome Analysis of Peach Fruits at Different Developmental Stages in Peach Kurakato Wase and Early-Ripening Mutant
    PENG JiaWei, ZHANG Ye, KOU DanDan, YANG Li, LIU XiaoFei, ZHANG XueYing, CHEN HaiJiang, TIAN Yi
    Scientia Agricultura Sinica. 2023, 56(5):  964-980.  doi:10.3864/j.issn.0578-1752.2023.05.012
    Abstract ( 88 )   HTML ( 15 )   PDF (3570KB) ( 44 )   Save
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    【Objective】 In this study, transcriptome analyses were carried out on the fruits of Kurakato Wase peach and its early-ripening mutant at different developmental stages. The key factors involved in fruit ripening regulation were explored, so as to provide a theoretical basis for further study on the regulation mechanism of fruit ripening. 【Method】 The flesh of Kurakato Wase peach and its early-ripening mutant was sampled at 30 d, 45 d, 59 d, 71 d, and 89 d after anthesis, and transcriptome analyses were performed on the above samples. The candidate differentially expressed genes (DEGs) were verified by quantitative real-time PCR (qRT-PCR). The biological function of DEGs were analyzed through GO function and KEGG pathway. The weighted gene co-expression network analysis (WGCNA) was constructed to identify the hub modules and hub genes closely related to fruit ripening. 【Result】 Four comparison groups including y1 vs c1, y2 vs c2, y3 vs c4 and y4 vs c5 were obtained based on fruit development stages. A tatal of 4 395 DEGs were identified with 2 212 up- and 2 183 down-regulated genes. There were 10, 11 and 18 candidate genes involved in ethylene, abscisic acid and auxin synthesis and signal transduction, respectively. The interaction networks between 10 IAA proteins and their predictive interacting proteins ARF were constructed. GO function revealed that the DEGs were mainly enriched in cellular processes, metabolic processes and monomeric processes in the biological process category; in cell component category, DEGs were mainly enriched in membranes and cellular components; in molecular function category, DEGs were mainly enriched in binding protein and catalytic activity. There were more DEGs in comparison groups y3 vs c4 and y4 vs c5, and these DEGs mainly enriched in molecular functions, such as binding and catalytic activity. The KEGG pathway analysis showed that a variety of secondary metabolites changed during fruit development and ripening, such as sesquiterpene and triterpenoid biosynthesis, flavonoid biosynthesis, carotenoid biosynthesis, and α-linolenic acid metabolism. In addition, auxin signal transduction pathway was found to be enriched at different time nodes. 【Conclusion】 Among DEGs, a large number of hormone signal transduction pathway genes, especially auxin signal pathway genes, were enriched, and these genes might play an extremely important role during fruit ripening. The functions of candidate genes IAA and ARF and the molecular regulation of fruit ripening would be further elucidated in the future studies.

    Lipid-Lowering Effects and Its Regulation Mechanism of Buckwheat Polyphenols in High-Fat Diet-Induced Obese Mice
    YAO YiJun, JU XingRong, WANG LiFeng
    Scientia Agricultura Sinica. 2023, 56(5):  981-994.  doi:10.3864/j.issn.0578-1752.2023.05.013
    Abstract ( 106 )   HTML ( 20 )   PDF (879KB) ( 31 )   Save
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    【Background】At present, relevant animal experiments have confirmed that buckwheat had a good intervention effect on obesity and lipid metabolism disorder caused by high-fat diet. At the same time, studies have shown that the adaptive thermogenesis of brown fat could effectively improve the body’s energy metabolism. Therefore, increasing the activity of brown fat and promoting the browning of white fat could be used as an effective way to prevent obesity and improve energy metabolism diseases.【Objective】The raw buckwheat and four kinds of main polyphenols identified from buckwheat were prepared according to the content proportion to study the lipid-lowering effects of buckwheat polyphenols in high-fat diet-induced obese mice via browning of white adipocytes.【Method】Phenolic compounds in buckwheat polyphenol extract were identified by UHPLC-Q-Orbitrap mass spectrometry combined with database. And the main polyphenol compound was prepared according to the content proportion in the raw buckwheat. In order to explore the regulating effects of buckwheat on body weight, organ index and blood lipid level in C57BL/6J mice, the dietary intervention was carried out with 10%, 20% and 40% proportion of buckwheat to replace basic feed and the 2.5 mg∙mL-1 of mixture buckwheat polyphenol standard substance. Last but not least, Western-blot and q-PCR were used to investigate the effects of buckwheat feed and polyphenol intervention on the expression of heat-producing proteins and genes in white and brown fat in mouse subcutaneous adipose tissue.【Result】Twenty phenolic compounds in buckwheat polyphenol extract were identified by UHPLC-Q-Orbitrap mass spectrometry, and the content of four phenolic compounds with the highest content (hydroxycinnamic acid, quercetin,scoparone, and rutin) accounted for more than 80% of the total phenolic compounds, which were the main polyphenols in buckwheat. The results showed that after 39 d in the diet intervention, compared with high-fat diet group, adding buckwheat feed and buckwheat polyphenol intervention on body weight in mice, the liver index and serum triglyceride (TG), total cholesterol (T-CHO), low-density lipoprotein cholesterol (LDL-c), aspartate aminotransferase (ALT) and third transaminase (AST) increased significantly inhibitory effect; with the increase of amount of buckwheat, the inhibition increased. At the same time, adding buckwheat feed and buckwheat polyphenol could protect and improve the decrease of renal index and HDL-c in mice. It was also the high dose 40% buckwheat and polyphenol group that had the best effect, which showed that phenolic substances played a major role in the improvement of abnormal lipid metabolism in high-fat diet-induced obese mice. UCP1, PRDM-16, and PGC-1α protein and gene expression levels were significantly decreased in the high-fat diet group compared with the normal diet group, while Tcf21 and HOXC8 expression levels were significantly increased (P<0.05). After dietary intervention, protein expression levels of UCP1, PRDM-16 and PGC-1α in the buckwheat intervention group were significantly up-regulated, while protein expression levels of Tcf21 and HOXC8 were significantly decreased (P<0.05). In addition, the 40% buckwheat and polyphenol intervention group significantly increased the expression of UCP1, PRDM-16 and PGC-1α genes in mouse visceral adipose tissue (P<0.05), and which down-regulated the expressions of Tcf21 and HOXC8 genes.【Conclusion】Buckwheat intervention could effectively improve the lipid metabolism disorder in high-fat diet-induced obese mice, and it was confirmed that buckwheat polyphenols played a major role. The mechanism was mainly buckwheat polyphenols could significantly improve the thermogenic activity of brown fat, reduce the content of white fat, and regulate the energy metabolism balance. This study provided a new idea and theoretical basis for the utilization of buckwheat and the development of functional foods about lipid-lowering products.

    Estimation of Genetic Parameters and Genome-Wide Association Study of Heat Indicators in Holstein Cattle Based on Collar-Mounted Device
    YANG MingLu, ZHANG HaiLiang, LUO HanPeng, HUANG XiXia, ZHANG HanLin, ZHANG ShiShi, WANG Yan, LIU Lin, GUO Gang, WANG YaChun
    Scientia Agricultura Sinica. 2023, 56(5):  995-1006.  doi:10.3864/j.issn.0578-1752.2023.05.014
    Abstract ( 129 )   HTML ( 31 )   PDF (1297KB) ( 62 )   Save
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    【Objective】 The study was to explore population characteristics and influencing factors of heat index and activity peak based on cattle collar data and system and to estimate its genetic parameters, and to identify the genetic markers related to the estrus indicators of cow, the genetic analysis and GWAS were performed for heat index and activity peak, so as to provide useful information for improving dairy cow fertility by formulating breeding strategies from the perspective of genetic breeding.【Method】The heat index and activity peak records of 2 074 milking cows from a large-scale dairy farm in Beijing area were collected from July 2017 to May 2020. The MIXED model in SAS 9.4 was used to analyze the impacts of various factors on heat index and activity peak in Holstein cattle, and the DMUAI module of DMU software was used to estimate the heritability and genetic correlations of heat index and activity peak. The GWAS for heat index and activity peak was performed based on De-regressed Proof (DRP) using FarmCPU software. 【Result】 In Chinese Holstein cattle, the average heat index was (71.77±19.13) au, and the activity peak was (105.94±30.73) au/2 h. The heritability of heat index and activity peak was 0.04±0.01 and 0.19±0.04, respectively. The genetic correlation between heat index and activity peak was 0.45±0.03. The approximate heat index between the age at first calving in heifer, interval from first to last insemination, and interval from first to last insemination was 0.37, -0.41, and -0.55, respectively. The GWAS for heat index and activity peak identified seven significant single nucleotide polymorphism loci (SNP) at the genome level, and 31 genes nearby the significant SNP (within 300 kb) were found. 【Conclusion】 The heat index and activity peak obtained by collar system were heritable, and there was a moderate genetic correlation between them. The activity peak was of low heritability and could reflect the heat index of cows to some extent. The activity peak and heat index provided a new direction for studying and breeding the estrus behavior of cows and improving the fecundity of cows. The function of PTGS1 (Prostaglandin-Endoperoxide Synthase1) and NDUFA8 (Ubiquinone Oxidoreductase Subunit A8) genes identified by GWAS was related to the total number of donor embryos, and the function of VEGFA (Vascular Endothelial Growth Factor A) gene was related to the reproduction of cows.

    Curcumin Alleviates Zearalenone-Induced Oxidative Damage in Porcine Renal Epithelial Cells via SIRT1/FOXO1 Pathway
    CUI HongJie, LU ChunTing, PAN LiQin, HU Hui, ZHONG PeiYun, ZHU JieYing, ZHANG KaiZhao, HUANG XiaoHong
    Scientia Agricultura Sinica. 2023, 56(5):  1007-1018.  doi:10.3864/j.issn.0578-1752.2023.05.015
    Abstract ( 123 )   HTML ( 19 )   PDF (2402KB) ( 55 )   Save
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    【Objective】 The purpose of this study was to investigate the protective effect of curcumin (Cur) on zearalenone (ZEA)-induced oxidative damage in porcine renal epithelial cells (PK-15), and to elucidate the protective mechanism based on SIRT1/FOXO1 signaling pathway. 【Method】 The experiment was divided into 5 groups: control group, ZEA group (36.55 μg·mL-1 ZEA), Cur6.25 group (36.55 μg·mL-1 ZEA+6.25 μmol·L-1 Cur), Cur12.5 group (36.55 μg·mL-1 ZEA+12.5 μmol·L-1 Cur), and Cur25 group (36.55 μg·mL-1 ZEA +25 μmol·L-1 Cur). MTT assay was used to determine the half inhibitory concentration of ZEA and the maximum safe concentration of Cur on PK-15 cells. The morphological changes were observed by inverted microscope. The levels of intracellular reactive oxygen species (ROS), superoxide dismutase (SOD), catalase (CAT), and malondialdehyde (MDA) were detected by the reagent kits. Real-time quantitative PCR was used to detect the mRNA levels of SIRT1, FOXO1, CAT and Mn-SOD. The expression levels of SIRT1, FOXO1 and Acetyl-FOXO1 proteins were detected by Western Blot. 【Result】 The IC50 of ZEA was 36.55 μg·mL-1, and the maximum safe concentration of Cur was 25 μmol·L-1. Compared with the control group, ZEA significantly decreased the cell viability of PK-15 cells (P<0.01), significantly increased the levels of ROS and MDA (P<0.01), and significantly decreased the activities of SOD and CAT (P<0.01). Compared with ZEA group, the different concentrations of Cur (6.25, 12.5, 25 μmol·L-1) significantly increased the cell viability of PK-15 cells (P<0.05) and improved the cell morphology. ROS and MDA levels induced by ZEA were also significantly reduced by Cur (P<0.01). Moreover, SOD and CAT activities in cells were significantly increased (P<0.01). qRT-PCR results showed that, compared with the control group, ZEA decreased SIRT1 mRNA expression, significantly increased FOXO1 mRNA expression (P<0.01), increased Mn-SOD mRNA expression, and significantly decreased CAT mRNA expression (P<0.01). Compared with ZEA group, mRNA expression levels of SIRT1 and CAT were increased in different degrees, FOXO1 mRNA expression levels were significantly decreased (P<0.01), and Mn-SOD mRNA expression levels were significantly increased (P<0.01) in all Cur groups. Western Blot results showed that ZEA significantly reduced SIRT1 protein expression (P<0.05), and significantly increased Acetyl-FOXO1 protein expression (P<0.01). Compared with ZEA group, SIRT1 protein expression was significantly increased (P<0.01), while Acetyl-FOXO1 protein expression was significantly decreased (P<0.01) in all Cur groups.【Conclusion】 Cur could up-regulate the expression of SIRT1, reduce the acetylation level of FOXO1, and induce the expression of antioxidant enzymes Mn-SOD and CAT, thereby eliminating ROS, reducing the level of MDA, and alleviating the oxidative damage of ZEA on PK-15 cells.