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    16 April 2024, Volume 57 Issue 8
    CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS
    Genetic Analysis and Candidate Gene Identification on Fertility and Inheritance of Hybrid Sterility of XI and GJ Cross
    XU Na, TANG Ying, XU ZhengJin, SUN Jian, XU Quan
    Scientia Agricultura Sinica. 2024, 57(8):  1417-1429.  doi:10.3864/j.issn.0578-1752.2024.08.001
    Abstract ( 22 )   HTML ( 3 )   PDF (4732KB) ( 8 )   Save
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    【Objective】The F1 hybrid sterility between XI/indica and GJ/japonica severely hinders the utilization of hybrid advantage between subspecies. Exploring the genetic mechanism and identifying new regulatory genes for XI/GJ hybrid sterility will provide theoretical basis for promoting genetic improvement of XI/GJ hybrid seed setting rate. 【Method】A series of stable genetic recombination inbred lines (RILs) containing 95 plant lines were derived from the cross between XI variety Habataki and GJ variety Sasanishiki after 10 generations inbred using single seed descent method. High throughput sequencing was performed on both parents and RILs on the Illumina platform, and the distribution of Habataki pedigree in RILs was analyzed at the whole genome level. The segregation distortion regions were identified, and hybrid sterile related gene loci were screened within the segregation distortion regions, then identified candidate genes through sequence alignment comparison. The targeted gene was knockout to verify the function using CRISPR gene editing technology. 【Result】The hybrid F1 plants derived from the cross between Habataki and Sasanishiki showed significant heterosis in panicles, grains per panicle, and thousand grain weight, but its seed setting rate significantly decreased. I2-KI microscopy revealed a significant decrease in F1 pollen fertility. High throughput sequencing of the entire genome of RILs revealed significant segregation distortion on Chr.1, Chr.3, Chr.5, Chr.6, Chr.7, and Chr.12, indicating that the genotype in this region tends towards the Habataki. Sequence alignment comparison revealed that Sc, S5, and HSA1 are target genes for the segregation distortion on Chr.3, Chr.6, and Chr.12. The CRISPR gene editing mutants with a knock-out Sc-Haba-3 allele in Habataki successfully improved the pollen fertility and seed setting rate of F1 hybrid with Sasanishiki. A complex structural variation was found between Sasanishiki and Habataki in the segregation distortion of Chr.1. A 24.7 kb segment containing 4 predicted genes in the Sasanishiki genome was replaced by a 64.8 kb segment containing 10 predicted genes in Habataki, the structural variation may involve in controlling the hybrid sterility of XI and GJ cross. 【Conclusion】This study detected multiple XI/GJ hybrid infertility related loci, and successfully improved F1 fertility by using CRISPR gene editing to knock out multiple copies of Sc in Habataki, locking in the target gene in the Sd region of Chr.1.

    Construction of SSR Fingerprint Library and Comprehensive Evaluation for Approved Cotton Varieties in China
    WU YuZhen, HUANG LongYu, ZHOU DaYun, HUANG YiWen, FU ShouYang, PENG Jun, KUANG Meng
    Scientia Agricultura Sinica. 2024, 57(8):  1430-1443.  doi:10.3864/j.issn.0578-1752.2024.08.002
    Abstract ( 20 )   HTML ( 1 )   PDF (2278KB) ( 7 )   Save
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    【Objective】Cotton, a heterotetraploid crop with a complex genome structure, faces challenges in achieving high homozygosity due to frequent cross-pollination. The absence of effective technical supervision in the cotton seed market and the persistence of disordered varieties have a negative impact on the consistency of fiber quality. The objectives of this study are threefold: to establish a DNA fingerprint database for approved cotton varieties in China over the past 20 years, to explore a high-throughput SSR identification model for cotton varieties, and to provide a basis for the authentication of existing varieties and the specific identification of new cotton varieties. Additionally, we aim to analyze the genetic diversity and population differentiation among approved varieties. Ultimately, our goal is to provide a theoretical framework for identifying cotton varieties that are well-suited to different ecological regions and for developing varieties that can adapt to new environments. 【Method】Based on multiplex PCR technology and capillary electrophoresis detection method, using 60 SSR markers screened to construct a DNA fingerprint library of 1 015 standard samples of cotton approved varieties. Through the plant variety DNA fingerprint library management system, the SSR fingerprints of approved varieties were compared pairwise to analyze the genetic differences of approved varieties and screen the core SSR loci for variety identification. Cluster analysis and population structure analysis were used to analyze the genetic diversity of 1 015 cotton approved varieties and calculate the genetic differentiation index between populations. 【Result】60 SSR markers amplified 216 allelic variations in 1 015 approved varieties, with an average of 3.6 allelic variations and a mean PIC value of 0.37. When the SSR fingerprints of the 1 015 approved varieties were compared, a total of 513 591 pairwise results were generated, with a maximum of 58 different loci between samples. The percentage of different loci was mainly concentrated at 41%-70%, involving 428 115 groups, accounting for 83.36%. Among them, when the percentage of different loci was at 51%-60%, the largest number of groups was involved, accounting for 197 829 groups, accounting for 38.52%. When the percentage of different loci between varieties was greater than 20%, it accounted for more than 99% of all pairwise comparison groups, and the pairwise comparison results with a percentage of different loci lower than 20% only accounted for 0.58%. Based on the combination identification method, a set of cores SSR loci containing 10 SSR loci was selected, and the discrimination ability among the 1 015 varieties reached 99%. Clustering results and population structure analysis showed that the 1 015 varieties were clearly divided into five subpopulations. G1 (n=240) was an early-maturing cotton subpopulation, mainly distributed in northern and inland regions of China. This subpopulation had the most abundant genetic diversity among varieties, with an average genetic distance of 0.419 between varieties. G2 (n=277) was a medium-maturing cotton subpopulation, distributed in the Yangtze River Basin. This subpopulation had more hybrids, with an average genetic distance of 0.309 within the subpopulation. G3 (n=109) belonged to early-maturing and medium-maturing cotton subpopulations, distributed in Hebei'sHeilonggang region. This subpopulation had relatively simple genetic components, with the smallest average genetic distance among upland cotton subpopulations at only 0.150. G4 (n=254) belonged to a medium-early maturing cotton subpopulation, mainly distributed in the Yellow River Basin. The average genetic distance within this subpopulation was 0.307. G5 (n=37) consisted of 37 sea island cotton samples, with the smallest average genetic distance within the population at only 0.149. The genetic differentiation level between sea island cotton and upland cotton was the highest, with an average FST value of 0.503. Among upland cotton populations, the genetic differentiation level between G3 and other subpopulations was the highest, with FST values ranging from 0.193 to 0.242. The genetic differentiation level between the Yangtze River Basin and the Yellow River Basin was the lowest, with an FST value of 0.112. 【Conclusion】A DNA fingerprint library of standard samples of 1 015 approved varieties in China over the past 20 years was constructed. A set of cores SSR loci containing 10 SSR loci was selected to clearly identify more than 99% of the varieties. A high-throughput cotton identification model of "core loci + extended loci" was created. The 1 015 varieties were divided into five subpopulations, and upland cotton had obvious geographical distribution characteristics.

    Effect of 14-Hydroxylated Brassinosteroids Growth Regulator on Growth and Yield of Rapeseed
    HE YongQiang, ZHANG JinKui, XU JinSong, DING XiaoYu, CHENG Yong, XU BenBo, ZHANG XueKun
    Scientia Agricultura Sinica. 2024, 57(8):  1444-1454.  doi:10.3864/j.issn.0578-1752.2024.08.003
    Abstract ( 14 )   HTML ( 1 )   PDF (740KB) ( 3 )   Save
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    【Objective】As a new Plant growth regulator, 14-hydroxylated brassinosteroids(14-HBR) increase biological activity by 50% than the traditional Brassicin sterols, while the relation effect of 14-HBR on rapeseed growth, yield and seed coating pesticide were not clear. 【Method】In this study, 14-HBR regulator and pesticide are used to treat the seeds in mid-duration winter rapeseed variety (Yangguang 2009) and early-duration variety (Yangguang 131), investigated germination, seedling growth, insect resistance, and yield, to reveal the interaction effects of environment, variety genotype and 14-HBR. 【Result】0.0075 and 0.015 mg·L-1 14-HBR treated seed increased significantly germination potential in medium duration rapeseed, but decreased germination potential and germination rate significantly as mixed with pesticide. 14-HBR treated seed had no significant effect on germination rate and germination potential in short duration rapeseed. The 14-HBR showed better biological activity in seedling growth and yield than that of Brassinolide, 0.0075 and 0.015 mg·L-1 Brassinolide increased by an average of 5.19% and 8.15%, 0.0075 and 0.015 mg·L-1 14-HBR increased by an average of 11.98% and 5.50%, respectively. 14-HBR mixed with seed pesticide of Clothianidin and Thiamethoxam, also increased seedlings weight and yield. The yield of Thiamethoxam and Clothianidin seed treatments increased by 4.7% and 4.6% independently. The yield of mixed with 0.0075 and 0.015 mg·L-1 14-HBR to Clothianidin increased 6.8% and 3.3%, mixed to Thiamethoxam increased by 3.5% and 8.2%, respectively. 14-HBR did not affect insecticidal activity of Thiamethoxam and Clothianidin to peach bud nymphs and phyllotreta striolata fabricius. 【Conclusion】The study revealed seed treatment with 14-HBR regulator has a positive effect on early growth of rapeseed and increased yield of rapeseed significantly, interacted with planting environment, pesticide type and varieties genotype as traditional regulators, it is necessary to optimize seed treatment technology of 14-HBR regulator to obtain higher harvest yield.

    TILLAGE & CULTIVATION·PHYSIOLOGY & BIOCHEMISTRY·AGRICULTURE INFORMATION TECHNOLOGY
    Effects of Postponing Nitrogen Fertilizer Application on Flag Leaf Physiological Characteristics and Yield of Spring Wheat Under High Temperature Stress
    LI YongFei, LI ZhanKui, ZHANG ZhanSheng, CHEN YongWei, KANG JianHong, WU HongLiang
    Scientia Agricultura Sinica. 2024, 57(8):  1455-1468.  doi:10.3864/j.issn.0578-1752.2024.08.004
    Abstract ( 19 )   HTML ( 2 )   PDF (687KB) ( 9 )   Save
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    【Objective】The effects of postponing nitrogen fertilizer on membrane lipid peroxidation, antioxidant enzyme activity, osmotic adjustment substance content in flag leaves and yield of spring wheat (Triticum aestivum L.) under high temperature stress were investigated, and the scientific fertilization methods to alleviate high temperature premature senescence of spring wheat after anthesis were screened. 【Method】From 2022 to 2023, Ning 3015 was used as the test material, and the test was carried out in the test base of the sixth team of Pingjipu Agricultural Reclamation in Ningxia. The split-plot experiment design was adopted. The main plot was temperature, with (25±2)℃ (normal temperature, RT) and (35±2)℃ (high temperature, HT), and the sub-plot was nitrogen fertilizer operation. The total amount of nitrogen application was constant, with 300 kg·N·hm-2: in 2022, G1 (30% of nitrogen application at tillering stage), G2 (30% of nitrogen application at jointing stage), G3 (30% of nitrogen application at booting stage), G4 (20% of nitrogen application at heading stage) and G5 (20% of nitrogen application at filling stage) were set up; in 2023, a total of three fertilization methods, including G1, G3 and G5, were selected from the previous year 's test results. Samples were taken every 5 days from the flowering stage to determine the indexes of membrane lipid peroxidation, antioxidant enzyme activity and osmotic adjustment substances in flag leaves. 【Result】Superoxide dismutase (SOD), peroxidase (POD), catalase (CAT) activities, proline (Pro) content in flag leaves and yield in two consecutive years of Nitrogen Fertilizer Transportation Experiment (NFTO) reached the highest under the G5 treatment, while malondialdehyde (MDA) and membrane permeability were the lowest under the G5 treatment, which were significantly different from the G1 treatment (P<0.01). According to Pearson's (Pearson) correlation analysis, it was shown that spring wheat grain yield was significantly positively correlated (P<0.05) with SOD, POD, CAT activities and proline content, while it was significantly negatively correlated (P<0.05) with MDA content and membrane permeability. Comprehensive evaluation showed that the ordering of the principal component scores of nitrogen fertilizer transport in 2022 and 2023 was G5>G3>G4>G2>G1 and G5>G3>G1, respectively, and the trend of the results of the two-year test was consistent. 【Conclusion】Appropriate nitrogen fertilizer postponing could reduce the membrane lipid peroxidation of spring wheat flag leaves under high temperature stress, increase the activity of antioxidant enzymes and the content of osmotic adjustment substances, and then increase the yield. In this experiment, the treatment of 20% total nitrogen application at filling stage was more suitable.

    Remote Sensing Detection of Cropping System and Its Spatial-Temporal Pattern in China
    ZHANG SuXin, SHEN Ge, YU QiangYi, WU WenBin
    Scientia Agricultura Sinica. 2024, 57(8):  1469-1489.  doi:10.3864/j.issn.0578-1752.2024.08.005
    Abstract ( 15 )   HTML ( 3 )   PDF (11925KB) ( 7 )   Save
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    【Objective】Cropping systems of cropland are the concrete embodiment of the mode of agricultural production, which reflect the coupled human-environment interactions. The formation is affected by natural resource elements and human land use behavior. This study aims to scientifically understand the spatial-temporal pattern of cropping systems, which helps to optimize agricultural distribution, improve agricultural production capacity, and realize the sustainable agriculture. 【Method】This study combined remote sensing monitoring with spatial decision tree models and other means to construct an inter-annual detection method system for cropping systems, which is designed for Chinese agricultural conditions, and then spatial pattern of cropping systems was analyzed. Firstly, the connotation of cropping systems was defined by identifying concepts such as cropping intensity, multiple cropping index, and considering of characteristics of “long-lasting” “periodicity” “stability”. Secondly, the indicators (i.e. continuity and frequency) were constructed, and were calculated at the pixel scale by the moving time window. Finally, the significance of the cropping intensity and characteristics of cropping system was evaluated. The decision tree method was also applied to determine the type of cropping systems, and the spatial-temporal heterogeneity of cropping systems in different regions was analyzed from the aspects of regional differences and dynamic laws. 【Result】 (1) Quantitatively, the largest area, 53.52%, is occupied by the single-cropping system, followed by the double-cropping system at 23.28%, the seasonal fallow system (i.e. 3 crops in 2 years) and the annual fallow system at 12.80% and 6.94%, respectively. (2) Spatially, the single-cropping system, double-cropping system, seasonal fallow system and annual fallow system are concentrated in Northeast China, North China, South of Yangtze River and “Sickle Bend” areas, respectively. (3) Temporally, it revealed the heterogeneity of cropping system and static multiple cropping index in the time dimension. For example, the regions with multiple cropping index of 1 in 2018 consist of 75.18% single-cropping system, 6.60% double-cropping system, 8.92% seasonal fallow system and 8.02% annual fallow system. 【Conclusion】This study proposed a method for mapping inter-annual cropping systems, combining remote sensing temporal monitoring and spatial decision tree models. It revealed the spatial pattern of cropping systems which is gathered by zone and cropping intensity is higher in the south and lower in the north. The Songnen Plain, “Sickle Bend” and other spatial gathering areas were intuitively displayed. Also, the differences between multi-cropping and cropping system were compared, which were mainly manifested in the spatial inconsistency between the cropping system and the annual multiple cropping index, as well as the periodicity of the cropping system. The results will provide case support for rationally increasing the cropland multi-cropping intensity and promoting the implementation of the “grain storage in the ground” strategy.

    PLANT PROTECTION
    The Transcription Factor NbMYB1R1 Inhibits Viral Infection by Promoting ROS Accumulation
    JIANG XingLin, YU LianWei, FU Han, AI Niu, CUI YingJun, LI HaoHai, XIA ZiHao, YUAN HongXia, LI HongLian, YANG Xue, SHI Yan
    Scientia Agricultura Sinica. 2024, 57(8):  1490-1505.  doi:10.3864/j.issn.0578-1752.2024.08.006
    Abstract ( 15 )   HTML ( 1 )   PDF (5329KB) ( 7 )   Save
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    【Background】Cucumber green mottle mosaic virus (CGMMV) is an important quarantine plant virus in China, which has seriously reduced the production of vegetables and melons worldwide. The MYB protein family is large, multifunctional and present in all eukaryotes. Most MYB proteins act as transcription factors that control development, and metabolism in plants, and regulate biotic and abiotic stress responses. Previous studies have shown that during CGMMV infection, the expression of transcription factor gene NbMYB1R1 was significantly up-regulated. 【Objective】The objective of this study is to clarify the mechanism of NbMYB1R1 involved in CGMMV infection, and to provide a theoretical basis for controlling CGMMV infection. 【Method】MEGA 7.0 was used to construct a phylogenetic tree to analyze the amino acid sequence of NbMYB1R1 protein. The expression vector NbMYB1R1-GFP was constructed, transformed into Agrobacterium GV3101 and infiltrated the tobacco leaves to observe the subcellular localization of NbMYB1R1 by confocal microscopy. The transcriptional levels of NbMYB1R1 at different stages of CGMMV infection and ROS related genes in silenced NbMYB1R1 plants were analyzed using qRT-PCR technology. TRV-mediated gene silencing (VIGS) was utilized to analyze the function of NbMYB1R1, NbAOX1a, and NbAOX1b during CGMMV infection. Transient overexpression of NbMYB1R1 and NbMYB1R1 mutant was conducted to analyze the effect of NbMYB1R1 on CGMMV infection. Trypan blue staining and DAB staining were used to observe whether the cell death caused by the transient overexpression of NbMYB1R1 was related to the programmed cell death (PCD) and the accumulation of ROS. Yeast two-hybrid system was used to verify whether NbMYB1R1 interacted with CGMMV related proteins. 【Result】Phylogenetic tree analysis showed that NbMYB1R1 belonged to the 1R MYB category and had high homology with MYB transcription factors in a variety of tobaccos. The results of subcellular localization showed that NbMYB1R1 was localized in the nucleus. In CGMMV-infected tobacco plants, the transcript level of NbMYB1R1 was significantly changed compared with healthy plants, which was significantly up-regulated at 8 and 12 d of CGMMV infection. CGMMV was inoculated into TRV:NbMYB1R1 and TRV:00 plants. The NbMYB1R1-silenced plants showed the mottled and curled symptoms at 3 dpi, while the control plant appeared the symptoms at 3.5 dpi. Silencing NbMYB1R1 could effectively promote the accumulation of CGMMV at mRNA and protein levels. The CGMMV accumulation in NbMYB1R1 and NbMYB1R1 mutant transiently overexpression leaves was detected, and the results showed that NbMYB1R1 overexpression could effectively inhibit CGMMV infection and DNA binding domain deletion mutant reduced the inhibition of CGMMV by NbMYB1R1. The results of trypan blue and DAB staining showed that NbMYB1R1 overexpression could induce the ROS accumulation and cell death. The transcription level of ROS-related genes in TRV:NbMYB1R1 and TRV:00 plant was also detected, and the results showed that silencing NbMYB1R1 could specifically up-regulate the transcription of AOX1a and AOX1b. After CGMMV inoculation on silencing endogenous genes NbAOX1a and NbAOX1b, the systemic leaves of silenced NbAOX1a and NbAOX1b plants showed mottled and curled symptoms at 4 dpi, while the control plants appeared symptoms at 3.5 d. Meanwhile, the results of CGMMV CP mRNA and protein levels also indicated that silencing NbAOX1a and NbAOX1b could effectively inhibit the accumulation of CGMMV. The yeast two-hybrid results showed that NbMYB1R1 did not directly interact with the CGMMV proteins. 【Conclusion】During the CGMMV infection, the defense-related gene NbMYB1R1 is up-regulated. It speculates that the upregulation of NbMYB1R1 inhibits the transcription of the downstream genes AOX1a and AOX1b and activates the production of ROS in cells, thereby inhibiting viral infection. However, NbMYB1R1 does not produce this effect through a direct interaction with the CGMMV viral protein. NbMYB1R1 plays an important role in CGMMV infection.

    Identification and Evaluation of Major Potato Cultivars Resistance to Globodera rostochiensis and Detection of Their H1 Resistance Gene Marker
    HUANG LiQiang, JIANG Ru, ZHU BoZhi, PENG Huan, XU Chong, SONG JiaXiong, CHEN Min, LI YongQing, HUANG WenKun, PENG DeLiang
    Scientia Agricultura Sinica. 2024, 57(8):  1506-1516.  doi:10.3864/j.issn.0578-1752.2024.08.007
    Abstract ( 15 )   HTML ( 1 )   PDF (1441KB) ( 4 )   Save
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    【Objective】Potato cyst nematodes (PCN), including Globodera rostochiensis and G. pallida, are damaging soilborne quarantine parasitic pests on potatoes. The G. rostochiensis has now distributed in seven counties of Yunnan, Guizhou, and Sichuan provinces in China, and several seed potato bases are threatened by the spread of G. rostochiensis. This study aimed to identify the resistance level of major potato cultivars to G. rostochiensis, clarify the presence of a known H1 resistance gene, and to provide a reference for screening and promoting resistant cultivars for controlling G. rostochiensis. 【Method】For bioassay, 15 major potato cultivars from the southwest mixed-cropping zone were inoculated with G. rostochiensis populations in an isolation greenhouse. The number of cysts per plant was counted and relative susceptibility was calculated, and finally the resistance of the cultivars was ranked according to a standard scoring notation. For H1 resistance gene identification, two molecular markers 57R and TG689, linked to the H1 for resistance to G. rostochiensis Ro1 pathotype, were used to identify 15 cultivars of the bioassay, with the BCH marker for amplification of the beta carotene hydroxylase gene as a control. For the field trial, a comparison of resistant and susceptible potato cultivars was conducted in a naturally infested field in 2020 and 2021 in Zhaotong, Yunnan Province. To estimate the initial (Pi) and final (Pf) populations of G. rostochiensis, soil sample was taken before planting and after harvested, and the reproduction factor (Pf/Pi) was calculated. The height of above-ground part of the plant at budding and initial flowering stages and the total yields at harvesting stage were measured. 【Result】Among the 15 major potato cultivars, five cultivars (Yunshu 505, Xuanshu 5, Huishu 15, Huishu 19, and Yunshu 304) are highly resistant. It’s hard for G. rostochiensis to reproduce on these cultivars. Two cultivars (Lishu 6 and Xuanshu 6) are moderately susceptible. Eight cultivars are highly susceptible, especially the Pf/Pi of Hui-2, Lishu 15 and Xuanshu 8 was higher than that of the susceptible control Huishu 16 (Pf/Pi=17.15). Additionally, the results of the two H1 gene-linked molecular markers were generally consistent, five cultivars (Yunshu 505, Xuanshu 5, Huishu 15, Yunshu 304, and Xuanshu 6) contain the H1. G. rostochiensis reproduction factor differed significantly on resistant and susceptible cultivars in the field. The average Pf/Pi values of highly resistant cultivars (with resistance score 9) were <1 in 2021 (0.04-0.12) and 2022 (0.05-0.14), indicating that nematode population densities decreased after planting of highly resistant cultivars. On the contrary, the average Pf/Pi values of the highly susceptible cultivars were >1.00 in 2021 (1.18-2.75) and 2022 (1.76-3.24), indicating an increase in nematode populations in the field after planting of highly susceptible cultivars. There were significant differences between cultivars in plant height and yield (P<0.05). The mean height of the five highly resistant cultivars was significantly higher than that of the eight highly susceptible cultivars in the two years. The cultivars Xuanshu 5, Huishu 15 and Huishu 19 had the highest yields of 51.67-56.48 and 33.28-40.57 t·hm-2 in two years, while Hui-2 had the lowest yields. 【Conclusion】The resistant cultivars in the southwest mixed-cropping zone show excellent resistance to G. rostochiensis, mainly carrying the H1, and can reduce G. rostochiensis population densities in the field. High yields are produced by the resistant cultivars Xuanshu 5, Huishu 15, and Huishu 19, while Yunshu 304 is utilized as a processing potato for zinc-rich crisps. More focused identification of resistance levels of local cultivars based on G. rostochiensis incidence is urgently required.

    SOIL & FERTILIZER·WATER-SAVING IRRIGATION·AGROECOLOGY & ENVIRONMENT
    Spatiotemporal Distribution Characteristics and Influencing Factors of Soil Inorganic Carbon in Shaanxi Province
    FENG XiaoLin, ZHANG ChuTian, XU ChenYang, GENG ZengChao, HU FeiNan, DU Wei
    Scientia Agricultura Sinica. 2024, 57(8):  1517-1532.  doi:10.3864/j.issn.0578-1752.2024.08.008
    Abstract ( 15 )   HTML ( 1 )   PDF (606KB) ( 6 )   Save
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    【Objective】Soil inorganic carbon (SIC) plays an important role in regulating global carbon cycle. However, the distribution characteristics and influencing factors of SIC at regional scales are not clear. The study on the temporal and spatial distribution of SIC and its key influencing factors in Shaanxi Province can provide the reference and basis for clarifying the role and status of inorganic carbon in the terrestrial ecosystem carbon cycle. 【Method】This study collected 65 and 142 soil samples from the 1980s and 2010s in Shaanxi Province, along with relevant data on geographical factors, climatic conditions, land use types, vegetation status and soil properties. Variance analysis and Random Forest (RF) model were used to analyze the temporal and spatial distribution characteristics of SIC content. The influencing factors of SIC content in Shaanxi Province were also discussed. 【Result】SIC content in the 1980s of Shaanxi Province was in the order of Northern Shaanxi > Guanzhong of Shaanxi > Southern Shaanxi. Compared with the 1980s, SIC content in Northern Shaanxi Province was decreased by 31.5% in 2010s, while it remained almost unchanged in Guanzhong of Shaanxi Province, which increased slightly in southern Shaanxi Province. From the 1980s to 2010s, the decrease of inorganic carbon content in different soil layers in 0-100 cm section ranged from 20.6% to 27.7%, with the greatest decreases in 0-20 cm and 80-100 cm soil layers. Random Forest model analysis showed that average annual rainfall, bulk density and pH were the top three most important factors affecting SIC content in both 1980s and 2010s, and SIC content was the highest when the average annual rainfall were 450-650 mm. Soil inorganic carbon content increased with the increase of pH. The inorganic carbon content of soil with low bulk density was higher than that of soil with high bulk density. 【Conclusion】In general, SIC content in Shaanxi Province showed a decreasing trend from north to south. Compared with the 1980s, SIC content in topsoil of Shannxi Province and also the whole soil profile of northern Shaanxi Province decreased significantly in the 2010s. The SIC content in the 1980s and 2010s were mainly influenced by average annual rainfall, pH and bulk density.

    Long-Term Sphagnum Cultivation in Cold Waterlogged Paddy Fields Increases Organic Carbon Content and Decreases Soil Extracellular Enzyme Activities
    GAO YaFei, ZHAO YuanBo, XU Lin, SUN JiaYue, XIA YuXuan, XUE Dan, WU HaiWen, NING Hang, WU AnChi, WU Lin
    Scientia Agricultura Sinica. 2024, 57(8):  1533-1546.  doi:10.3864/j.issn.0578-1752.2024.08.009
    Abstract ( 16 )   HTML ( 1 )   PDF (4065KB) ( 4 )   Save
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    【Objective】In southern China, cold waterlogged paddy fields cover an expansive area and hold considerable potential for carbon sequestration and remittance. However, the low yield and modest income derived from rice cultivation in such paddy fields have led to a high rate of abandonment. This study investigated whether conversion of cold waterlogged paddy fields to Sphagnum cultivation, an economically important plant beneficial for carbon sequestration, significantly enhances soil carbon storage while improving the income of farmers. The overall aim of this study was to evaluate the impact of Sphagnum cultivation on the carbon sequestration potential of cold waterlogged paddy soils. 【Method】Zilinshan Village, Dushan County, Qiannan Prefecture, Guizhou Province, was selected as the study site. The physicochemical properties, extracellular enzyme activities, and organic carbon content in the surface soil (0-10 cm depth) were analyzed after transformation of cold waterlogged paddy fields to Sphagnum cultivation for 1, 3, 10, and 20 years. Paddy fields growing rice were used as the control. 【Result】 (1) The years of Sphagnum cultivation altered the physicochemical properties of cold waterlogged paddy soils. Especially after Sphagnum cultivation for 10 years, the soil bulk density, mean weight diameter of aggregates, and total phenol content were increased by 16.9%, 33.8%, and 88.1%, respectively, compared with the control. (2) With an increase in years of Sphagnum cultivation, the activities of cellulose hydrolase, acid phosphatase, β-1,4-N-acetylglucosaminidase, β-1,4-glucosidase, leucine aminopeptidase, and polyphenol oxidase significantly decreased. (3) After Sphagnum cultivation for 10 years, soil organic carbon and recalcitrant organic carbon contents increased significantly, and the dissolved organic carbon and easily oxidizable carbon contents decreased significantly, the activities of cellulose hydrolase, acid phosphatase, β-1,4-N-acetylglucosaminidase, β-1,4-glucosidase, leucine aminopeptidase, and polyphenol oxidase significantly decreased after Sphagnum cultivation for 20 years. (4) The structural equation model revealed that the years of Sphagnum cultivation had maximum direct positive effect on soil organic carbon and recalcitrant organic carbon. In terms of dissolved organic carbon and easily oxidizable carbon, they directly influenced by extracellular enzyme activity to the greatest extent. Generally, soil physicochemical properties have indirect effects on the four kinds of carbon through extracellular enzyme activities,, and the years of Sphagnum cultivation indirectly influenced four types of carbon through soil physicochemical properties. 【Conclusion】 The planting of Sphagnum moss can induce changes in the soil environment, leading to a significant increase in soil organic carbon content and a reduction in extracellular enzyme activity in waterlogged paddy fields. Additionally, it promotes carbon accumulation, with long-term Sphagnum planting further enhancing this process.

    Effect of Phosphorus Fertilizer Application Rates on the Loss of Colloidal Phosphorus on Purple Soil Slopes
    ZHONG JinPing, ZHENG ZiCheng, LI TingXuan, HE XiaoLing
    Scientia Agricultura Sinica. 2024, 57(8):  1547-1559.  doi:10.3864/j.issn.0578-1752.2024.08.010
    Abstract ( 12 )   HTML ( 1 )   PDF (690KB) ( 3 )   Save
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    【Objective】The risk of phosphorus loss from farmland is closely related to the amount of phosphorus fertilizer. Given the important role of soil colloids in the process of phosphorus transport and transformation at the soil-water interface, the effect of phosphorus fertilizer application on the loss of phosphorus from colloidal state on purple soil slopes and its relationship with runoff and sand production were explored, in order to provide the scientific basis for the understanding of phosphorus transport mechanism from the soil colloid point of view. 【Method】 Combining artificial simulated rainfall with laboratory analysis, the experiment was conducted to study the characteristics of abortion sediment production and colloid phosphorus loss on purple soil slope under the dosage of phosphorus fertilizer 0 (P0), 20 (P20), 40 (P40) and 100 (P100) mg·kg-1. 【Result】 Surface runoff was less affected by phosphorus fertilizer application, and erosion sand production was more affected by phosphorus fertilizer application. The initial sand production of the slope was significantly reduced by 49.3%-68.7% after phosphorus application, and the cumulative sand production was significantly reduced by 26.5%-30.9% under P100 treatment compared to the other phosphorus treatments. Surface runoff was the main loss pathway of water-dispersible total phosphorus (WTP) and colloidal phosphorus (CP) from purple soil slopes, which accounted for 57.5%-93.9 and 62.3%-94.8% of the total loss, respectively; CP was the main form of WTP loss from surface runoff, which accounted for 72.1%-80.7% of the WTP loss. Phosphorus application significantly increased the risk of phosphorus loss. Compared with P0 treatment, the cumulative loss loads of surface runoff WTP, CP, and DP (dissolved phosphorus) under phosphorus fertilizer application treatments were increased by 2.56-20.97, 2.72-22.21, and 1.17-10.40 times after phosphorus application, respectively, and the cumulative loss loads of eroded sediment WTP, CP, and DP were increased by 0.24-0.92 times, 0.05-1.09 times, 0.47-0.76 times, respectively. 【Conclusion】 The main pathway of colloidal phosphorus loss from purple soil slopes was surface runoff, and the characteristics of concentration change were closely related to the flow production process, while the loss load mainly depended on the phosphorus content of slope soil and the amount of phosphorus fertilizer applied. Total water dispersible phosphorus and colloidal phosphorus showed a highly significant correlation, colloidal phosphorus was the main form of phosphorus loss on purple soil slopes, and CP loss on slopes could be reduced by regulating surface runoff and reducing the amount of phosphorus fertilizer.

    HORTICULTURE
    Evaluation of Fruit Texture Quality in Melon
    YANG YaHeng, JIA PeiLong, NIE LanChun, ZHAO WenSheng, ZHAO JiaTeng, WANG JinXiang, LIU Jie
    Scientia Agricultura Sinica. 2024, 57(8):  1560-1574.  doi:10.3864/j.issn.0578-1752.2024.08.011
    Abstract ( 14 )   HTML ( 3 )   PDF (1398KB) ( 5 )   Save
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    【Objective】The aim of this study was to evaluate the fruit texture of melon (Cucumis melo L. subsp. melo) germplasm resources, so as to provide reference and basis for establishing fruit texture evaluation standards and breeding excellent varieties of melon. 【Method】Texture profile analysis (TPA) (including TPA hardness, springiness, chewiness, cohesiveness and resilience), puncture test (PT) (including puncture hardness, crispness and adhesiveness) and sensory evaluation method (sensory hardness, sensory crispness, juiciness, compactness and pulp texture quality) were performed to evaluate the texture of 278 melon germplasms. Through correlation analysis, stepwise regression analysis and factor analysis, the relationship model between sensory evaluation indexes and texture analyzer test indexes were established, and the important indexes of fruit texture evaluation in melon were defined. 278 melon germplasm resources were classified according to the indexes of fruit texture. 【Result】There was a significant correlation between texture analyzer test indexes and sensory evaluation indexes of fruits texture in melon. The prediction models of sensory hardness, sensory crispness, juiciness, compactness and pulp texture quality were established relying on the independent variables of hardness, crispness, adhesiveness, springiness, chewiness, cohesiveness and resilience evaluated by the texture analyzer, respectively. Three common factors were selected from the factor analysis of texture analyzer test indexes, and the cumulative variance contribution rate was 89.377%. The first common factor reflected the chewiness of pulp, the second common factor reflected the adhesiveness of pulp, and the third common factor reflects the resilience of pulp. Chewiness, adhesiveness and resilience were important parameters affecting the fruit texture of melon. According to the texture indexes, 278 melon were divided into three groups, and each group could be divided into two subcategories. The group I was characterized by the highest hardness, crispness and compactness, the coarsest pulp and the lowest juiciness. The group III was characterized by the lowest hardness, crispness and compactness, the thinnest pulp and the highest juiciness. The group II was characterized by an intermediate level of each index. 【Conclusion】Texture analyzer test indexes could reflect the texture quality of melon fruits. Chewiness, adhesiveness and resilience were important indexes to evaluate the melon fruit texture.

    Transcriptome and Proteome Association Analysis to Revealthe Molecular Mechanism of Baxi Banana Seedlings in Response to Low Temperature
    LIN Wei, WU ShuiJin, LI YueSen
    Scientia Agricultura Sinica. 2024, 57(8):  1575-1591.  doi:10.3864/j.issn.0578-1752.2024.08.012
    Abstract ( 15 )   HTML ( 2 )   PDF (6769KB) ( 4 )   Save
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    【Objective】 Low temperature is a significant natural disaster that affects banana production. In this study, based on transcriptome and proteome association analysis, the regulatory network of genes, proteins, signals and metabolic pathways involved in banana cold resistance was investigated. The aim was to explore the molecular mechanism of banana cold resistance. 【Methods】 ‘Baxi’ banana (Musa nana Lour) was treated at 7 ℃ for 1 and 3 d, and a control group was treated at 28 ℃. Based on the proteome data obtained in the previous study, the transcriptome sequencing technology was used to detect changes in the gene regulatory network of banana under cold stress. Simultaneously, the correlation analysis was conducted with proteomics to analyze the molecular mechanism of banana response to cold stress. 【Result】 Transcriptome analysis revealed that 11 370, 15 460 and 9 619 differentially expressed genes were identified in the three comparison groups of Cold1 vs CK, Cold3 vs CK and Cold1 vs Cold3, respectively. KEGG enrichment analysis of these genes revealed that the differentially expressed genes were enriched in several key signaling metabolic pathways, such as photosynthesis signal, glutathione metabolism, α-linolenic acid metabolic pathway and phenylpropanoid biosynthesis under low temperature stress. Moreover, there were significant differences in the enrichment degree of glutathione metabolic pathway between Cold1 d vs CK and Cold3 d vs CK. Real-time quantitative PCR analysis (qRT-PCR) was performed on several differentially expressed genes. Among them, the expression levels of key low-temperature regulatory genes, such as DREB, MAPK and MYB, were significantly increased after the low-temperature treatment. The expression trend of the selected 20 genes was essentially consistent with that of RNA-seq, confirming the accuracy of RNA-seq. The results of the transcriptome and proteome association analysis showed a positive correlation between the transcriptome and proteomics. A total of 6 211 proteins corresponding to transcripts were identified. Among these, 105 transcripts and their proteins were up-regulated, while 218 transcripts and their proteins were down-regulated. GO enrichment analysis showed that the differentially expressed genes and proteins were enriched in functions, such as photoresponse, chloroplast and oxidoreductase activity. Furthermore, the correlation analysis of differentially expressed genes and protein KEGG pathway revealed that the low temperature treatment suppressed the expression of genes and proteins related to phenylpropanoid biosynthesis and photosynthesis signaling pathway, while promoting the expression of proteins associated with α-linolenic acid metabolism and the glutathione pathway. 【Conclusion】 The transcriptome and proteomics were used to map the regulatory network of banana cold resistance at the gene and protein levels. It was found that the signal pathway of banana response to low temperature mainly involved photosynthesis signal, glutathione metabolism, α-linolenic acid metabolism and phenylpropanol biosynthesis.

    FOOD SCIENCE AND ENGINEERING
    The Stabilization of Aroma and Color During Hutai-8 Rose Winemaking by Gallic Acid Treatment
    FENG Fan, JIANG XingRui, WANG LingYun, ZHANG YongGang, LI AiHua, TAO YongSheng
    Scientia Agricultura Sinica. 2024, 57(8):  1592-1605.  doi:10.3864/j.issn.0578-1752.2024.08.013
    Abstract ( 15 )   HTML ( 3 )   PDF (1713KB) ( 6 )   Save
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    【Objective】This study was aimed to investigate the effects of gallic acid treatment on color and aroma preservation during the aging process of Hutai-8 rosé wine, in order to optimize the design of color and aroma enhancement techniques for rosé wine production. 【Method】In this study, Hutai-8 grape was used as raw material. Gallic acid was added at three different stages, including pre-fermentation (Pr), mid-fermentation (M), and post-fermentation (Po), with the concentrations of 200 and 300 mg∙L-1. After a 6-month storage period following fermentation, the aroma compounds of the wine samples were analyzed by headspace solid-phase microextraction-Gaschromatography-mass spectrometry (HS-SPME-GC-MS). The color parameters (L*, a*, b*, C*ab, Hab, and Δ*Eab) were determined by the CIELab color space parameters, and the color indices were measured by ultraviolet spectrophotometer (UV). Finally, the sensory evaluation was conducted to analyze the influences of different treatments on the aroma characteristics of the wine samples. 【Result】The post-fermentation treatments significantly increased the content of fermentation aroma compounds compared with CK, with an increase of approximately 16%. However, there was little difference between treatments of the two additive concentrations. The pre-fermentation treatments positively contributed to the preservation of varietal aroma compounds, with an increase of approximately 65%-73% compared with CK. The mid-fermentation treatments had a lesser stabilizing effect on the aroma compounds of the wine. Sensory evaluation results showed that post-fermentation treatment had the best effect on improving the overall aroma of the wine, and the pre-fermentation treatment was more effective than the mid-fermentation treatment. PLSR models revealed that terpenols, fatty acids, higher alcohols, acetates and fatty acid ethyl esters were the main aroma contributors (regression coefficients>0.1) to floral and citrus attributes (R2c>0.80 & R2v>0.70), with fatty acid ethyl esters and acetates being particularly prominent contributors. The color analysis results showed that pre-fermentation had a significant color-preserving effect, and the treatment with 200 mg∙L-1 (Pr1) was more effective than the treatment with 300 mg∙L-1 (Pr2). Comparied with CK, Pr1 treatment group’s, L* value decreased by 0.58% and a* value increased by 45.38%. Furthermore, the color characterization results showed that pre-fermentation treatment enhanced the purple-red tone of Hutai-8 rosé wine and the treatment with 200 mg∙L-1 was more effective than the treatment with 300 mg∙L-1. 【Conclusion】The addition of 200 mg∙L-1 gallic acid before fermentation had a significant effect on stabilizing the color of Hutai-8 rose wine, while the post-fermentation treatments could significantly increase the fermentative aroma content of wine.

    ANIMAL SCIENCE·VETERINARY SCIENCE
    lincRNA Cox2 Regulates BCG-infected Macrophages Glycolysis by miR-129-5p/AMPK
    XU Lei, YU JiaLin, LIU Li, DENG GuangCun, WU XiaoLing
    Scientia Agricultura Sinica. 2024, 57(8):  1606-1619.  doi:10.3864/j.issn.0578-1752.2024.08.014
    Abstract ( 12 )   HTML ( 1 )   PDF (3774KB) ( 3 )   Save
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    【Objective】 The aim of this study was to investigate the regulatory role of lincRNA Cox2 in the glycolysis of RAW264.7 macrophages infected by Bacillus Calmette-Guerin (BCG), and to elucidate the interaction between Mtb and macrophages, so as to provide a new target for the diagnosis and treatment of tuberculosis. 【Method】 RNA interference technique was used to knock down the expression of lincRNA Cox2, and miR-129-5p mimics were used to overexpress miR-129-5p. QPCR was performed to measure the lincRNA Cox2, miR-129-5p and proinflammatory cytokine ( IL-1β, TNF-α, and IL-6 ) expression after BCG infection. The expression of Lactic Acid was detected by Lactic Acid assay kit. The bacterial load was measured bacterial load in BCG-infected macrophages. Dual luciferase reporter gene system validation experiments were carried out on lincRNA Cox2 and miR-129-5p, or miR-129-5p and AMPK relationships. The expression of AMPK (AMP activated protein kinase), HK1 (Hexokinase 1), PKM2 (pyruvate kinase M2), and LDHA (Lactate dehydrogenase A) were detected by Western blotting. 【Result】 The expression of lincRNA Cox2 was significantly upregulated (P=0.000013) after BCG infection in RAW264.7 macrophages for 12 h. Compared with the BCG group, the siRNA+BCG group had significantly upregulated the expression of AMPK (P=0.000771), HK1 (P=0.00323), PKM2 (P=0.000135), LDHA (P=0.002532), and the secretion of LD (P=0.020802), but the expression of IL-1β (P=0.000451), TNF-α (P=0.000147), IL-6 (P=0.0001) was significantly reduced. The lincRNA Cox2 knockdown caused a significant reduce of bacterial load in BCG-infected macrophages (P=0.000127). Dual luciferase reporter gene system were performed to the co-localized of lincRNA Cox2 and miR-129-5p, and targeting AMPK. The expression of miR-129-5p was significantly reduced (P=0.000156) after BCG infection in RAW264.7 macrophages for 12 h. Compared with the BCG group, the miR-129-5p mimics+BCG group had significantly reduced the expression of AMPK (P=0.000262), HK1 (P=0.019524), PKM2 (P=0.001658), LDHA (P=0.000887), and the secretion of LD (P=0.044952). 【Conclusion】 lincRNA Cox2 promoted BCG-infected RAW264.7 macrophages glycolysis process by sponging miR-129-5p and targeting AMPK.

    Preparation and Application of Polyclonal Antibodies Against Pig CD1d Protein
    LIU ChuanXia, CHEN Xin, WANG Xiao, LI XueWen, LI TingTing, WENG ChangJiang, ZHENG Jun
    Scientia Agricultura Sinica. 2024, 57(8):  1620-1628.  doi:10.3864/j.issn.0578-1752.2024.08.015
    Abstract ( 11 )   HTML ( 1 )   PDF (1609KB) ( 3 )   Save
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    【Objective】 The aim of this study was to prepare polyclonal antibodies against porcine CD1d protein, so as to lay the foundation for exploring the function of porcine CD1d protein in the process of African swine fever virus (ASFV) infection. 【Method】 In this study, the pig CD1d gene was amplified using PCR and homologously recombined into the pGEX-6p1 vector, constructing a prokaryotic recombinant expression plasmid pGEX-6p1-CD1d. The recombinant plasmid E. coli BL21 (DE3) was transformed and induced for expression using IPTG. The expressed GST CD1d recombinant protein was identified by SDS-PAGE and Western blot (WB) methods. The SDS-PAGE results showed a clear band at approximately 50 ku, which was expressed in the form of an inclusion body. Then, protein purification was performed using glutathione agarose affinity chromatography. The purified GST-CD1d protein was mixed and emulsified with an equal volume of Freund's complete adjuvant. The purified protein was immunized in New Zealand white rabbits and administered subcutaneously at multiple points on the neck and back, with an immune dose of 200 μG/piece, and then second and third immunizations were performed at the 3rd and 5th weeks after the first immunization, respectively, using Freund's incomplete adjuvant emulsification, with the same method and dosage as the first immunization. On the 7th day after the third immunization, the blood was collected from the ear vein to isolate the serum. The fourth immunization was conducted at the 7 weeks after the first immunization, and the blood was collected from the heart one week later. The antibody was purified by Protein G affinity chromatography and frozen at -80 ℃. The expression and cellular localization of endogenous CD1d protein expressed by transient transfection of exogenous and porcine primary macrophages (PAMs) were indentified by using WB and indirect immunofluorescence (IFA). Similarly, the prepared CD1d antibody could pull down CD1d expressed by transient exogenous transfection through IP. In order to investigate the early stage of ASFV infection, ASFV was inoculated into PAMs and samples of ASFV infection for 0, 15, 30, and 60 minutes were prepared, respectively. CD1d was used as the primary antibody and the expression of CD1d protein was detected by WB. Plasmids pCAGGS-HA-CD1d and pCAGGS-Flag-CD2v were co transfected into HEK293T cells. After 24 hours, the cells were collected for lysis, and Flag beads overnight binding protein was added. The interaction was detected by WB staining. At the same time, the plasmids were cotransfected into HEK293T cells in a confocal dish, incubated with labeled antibodies, and corresponding fluorescent secondary antibodies were selected. The co localization of CD1d and CD2v was observed under a laser confocal microscope. Verification of Co-IP interaction between CD1d and ASFV outer capsule protein CD2v was verified. 【Result】 The GST-CD1d protein expressed in prokaryotic cells was expressed in the form of inclusion bodies, with a molecular weight of approximately 35 ku; After four rounds of immunization with CD1d recombinant protein in experimental rabbits, blood was collected and serum was separated. The purified antibody was detected by SDS-PAGE and showed a specific band at 45 and 25 ku, respectively, representing the heavy and light chains of the CD1d antibody. The rabbit anti CD1d antibody prepared using purified CD1d protein as immunogen contained both heavy and light chains, and had good purity; This antibody could identify the expression and cellular localization of transient transfected exogenous and PAMs endogenous CD1d proteins through WB and IFA. Further testing results showed that after ASFV infection with PAMs, the expression level of CD1d protein significantly increased, and WB and IFA results showed that CD1d interacted and co localized with the outer capsule protein CD2v encoded by ASFV. 【Conclusion】 This study prepared antibodies against CD1d through prokaryotic expression technology, laying the foundation for further exploration of the biological function of CD1d protein in ASFV infection process.