Scientia Agricultura Sinica

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Scientia Agricultura Sinica. 2021, 54(4): 0.

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Analysis of Differential Metabolites in Grains of Rice Cultivar Changbai 10 Under Salt Stress

ZHANG GuiYun,ZHU JingWen,SUN MingFa,YAN GuoHong,LIU Kai,WAN BaiJie,DAI JinYing,ZHU GuoYong

Scientia Agricultura Sinica. 2021, 54(4): 675-683. doi:10.3864/j.issn.0578-1752.2021.04.001

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【Objective】To reveal the influence of salt stress on metabolites and metabolism pathway in the rice seed of Changbai 10, understand the mechanism of its salt tolerance, and screen possible markers for salt tolerant rice varieties. 【Method】Gas chromatography-mass spectrometry (GC-MS) was used to analyze the changes of metabolites in the seeds (25 days after flower) of a saline tolerance japonica rice Changbai 10 under salt stress (4‰). The rice variety Yandao 8 with moderate salt tolerance was treated with the metabolite which was the highest improvement among the different metabolites to identify whether it could improve the salt tolerance. The treatment methods were as follows: Yandao 8 seeds were soaked in NaCl solution (5‰) containing the metabolite with different concentration gradient (0-400 mg·L^-1), after 20 hours, the seeds were put in germination box for germination test.【Result】A total of 295 metabolites were detected in the samples. Principal component analysis (PCA) and orthogonal partial least-squares-discriminant analysis OPLS-DA were performed to visualize the metabolic difference among experimental groups. The results revealed a total of 71 metabolites were significantly changed (42 up-regulated and 29 down-regulated) under salt stress. The top ten up-regulated metabolites were 5-hydroxynorvaline (5.23 fold), Pentane-1,2,5-triol (4.10 fold), Cyanoalanine (2.97 fold), L-asparagine (2.91 fold), 2-hydroxybutanoic acid (2.86 fold), Oxoglutaric acid (2.74 fold), Pyruvic acid (2.50 fold), Ornithine (2.40 fold), Citrulline (2.07 fold), Proline (1.94 fold). And the top ten down-regulated metabolites were Erythrose (2.83 fold), Xylonolactone (2.49 fold), Aspartate (2.42 fold), Uridine 5’-monophosphate (2.37 fold), Cholesterone (1.90 fold), Hexaric acid (1.87 fold), D7-glucose (1.86 fold), 5,7-dihydroxy-4’-methoxyisoflavone (1.72 fold), Amidosulfonic acid (1.61 fold), 6-deoxygalactofuranose (1.61 fold). Among them, 15 differential metabolites were KEGG annotated and significantly(P<0.01) affected arginine biosynthesis, alanine, aspartate and glutamate metabolism, butanoate metabolism, carbon metabolism, lysine degration, biosynthesis of amino acids, glyoxylate and dicarboxylate metabolism, citrate cycle (TCA cycle) and so on. The germination rate of Yandao 8 was only 66.7% under salt stress (5‰), significantly lower than that with exogenous 5-hydroxynvaline treatment (74.3-90.3%). The result revealed that germination rate of the seeds with 200 mg·L^-1 5-hydroxynvaline treatment was the highest.【Conclusion】Amino acid and organics acid were mainly up regulated, while sugar and lipid were mainly down regulated in salt treated group. The changed metabolism pathway of Arginine biosynthesis and TCA cycle might contribute to its high salt tolerance. 5-hydroxynvaline could effectively alleviate the inhibition of salt stress on rice seeds germination, and the optimal concentration is 200 mg·L^-1.

Evaluation of Resistance to Stripe Rust and Molecular Detection of Resistance Gene(s) in 243 Common Wheat Landraces from the Yunnan Province

XI Ling, WANG YuQi, YANG Xiu, ZHU Wei, CHEN GuoYue, WANG Yi, QIN Peng, ZHOU YongHong, KANG HouYang

Scientia Agricultura Sinica. 2021, 54(4): 684-695. doi:10.3864/j.issn.0578-1752.2021.04.002

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【Objective】As one of the most destructive wheat diseases in the world, stripe rust is an airborne fungal disease caused by fungus Puccinia striiformis. f. sp. Tritici, (Pst). Breeding resistant cultivars by exploiting resistant genetic resources is the most economical and effective strategy to combat the rusts. To provide theoretical basis for the management of wheat stripe rust by identifying and evaluating the resistance level of Yunnan wheat landraces to the current predominant races of the pathogen, and synthetically screened resistance genes that may be carried in the tested materials. 【Method】In this study, 243 wheat landraces originated from Yunnan were inoculated with two highly toxic and prevalent stripe rust races CYR32 and CYR34 at seedling stage. At the adult plant stage of wheat, the mixture of the current predominant races of CYR32, CYR33, CYR34, and Guinong Pathogenic group was used as the inoculums to test the resistance of the tested wheat. The wheat landraces were screened with the molecular markers closely linked to known stripe rust resistance genes Yr5, Yr10, Yr15, Yr18, Yr26, Yr28, Yr29, Yr30, Yr36, Yr39, Yr41, Yr48, Yr65, Yr67, Yr80, and Yr81.【Result】The results showed that the 243 of the tested wheat materials, of which 18 were resistant to CYR32, 32 resistant to CYR34, and only 8 (3.29%) were resistant to both races at the seedling stage. Based on identification results of the seedling stage and the adult plant stage, 174 (71.6%) exhibited adult plant resistance, of which 105 were high resistance to stripe rust. The results of molecular detection indicated that 48, 44, 4, 6, 4, and 101 out of the 243 tested wheat materials carried the resistance genes Yr10, Yr18, Yr29, Yr30, Yr65, and Yr81, respectively. There were 34, 4 and, 1 simultaneously carrying 2, 3, and 4 resistance genes, respectively. Yr5, Yr15, Yr26, Yr28, Yr36, Yr39, Yr41, Yr48, Yr67, and Yr80 were not detectable in all of the wheat landraces. In addition, 74 landraces not detected any of the 16 resistance genes mentioned the above, and 58 exhibited adult plant resistance, which, presumably, may carry other known or new resistance genes to wheat stripe rust.【Conclusion】As a special wheat germplasm resource in China, Yunnan wheat landraces have excellent resistance to stripe rust. This study determined that resistance of the 243 tested wheat landraces to the current predominant races of the pathogen was high on the whole, and 174 wheat landraces with stable resistance were identified. Among them, 74 lines might carry other known or new resistance genes to wheat stripe rust, which could be used as parents’ sources for further exploration of new stripe rust resistance genes or QTL.

Using CRISPR/Cas9-mediated Targeted Mutagenesis of ZmFKF1 Delayed Flowering Time in Maize

YANG Min,XU HuaWei,WANG CuiLing,YANG Hu,WEI YueRong

Scientia Agricultura Sinica. 2021, 54(4): 696-707. doi:10.3864/j.issn.0578-1752.2021.04.003

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【Objective】FKF1 is a key gene that plays an important role in many plant flowering pathways. In order to study the function of FKF1 in maize, ZmFKF1 editing mutants were obtained by using CRISPR/cas9 technology. In this study, we used these mutants as materials to clarify the role of ZmFKF1 in maize flowering pathway through phenotypic analysis and the expression analysis of key flowering genes. The result provide a theoretical basis for molecular breeding and genetic improvement of maize.【Method】 ZmFKF1 gene was cloned from B104, and its structure was determined by sequence alignment. The targeted sequences of ZmFKF1 were designed according to the principle of CRISPR/Cas9 technology, then these targeted sequences were compared and analyzed in maize reference genome, and the non-specific target site was excluded. Finally, ZmFKF1-T1 on exon 1 of ZmFKF1 was selected to construct CRISPR/cas9 gene editing expression vector. At the same time, B104 immature embryos were selected as explants to transform by Agrobacterium-mediated genetic transformation, the resistant calli were obtained through resistance screening, and then buds and roots were induced. The T₀ generation ZmFKF1 gene editing positive plants were obtained and verified by using the specific primers of cas9. The target site amplification and sequencing analysis was used to determine whether the T₁ transgenic lines had mutation at the expected target site of ZmFKF1 and the type of mutation, and screened the homozygous lines of ZmFKF1 site mutation. After obtaining the above materials, the flowering phenotypes of these materials were statistically analyzed with wild type as control. At the same time, qRT-PCR was used to detect the expression of key genes that related to flowering pathway in the above materials and further verify the phenotype. 【Result】The target sequence was designed on exon1 of ZmFKF1 to construct gene editing expression vector. The transgenic lines were obtained by genetic transformation realized site directed mutagenesis of ZmFKF1. A total of 18 T₀ generation ZmFKF1 gene-edited lines were obtained, and 6 of them were generated mutation on the exon1 with two different mutations types, including single base insertion and multiple bases deletion. The phenotype analysis shown that the flowering time of three T₂ generation ZmFKF1 homozygous lines was delayed compared with wild type B104. Furthermore, the expression levels of three related flowering pathway genes (ZmGI, conz1 and ZmZCN8) in the mutants were significantly lower than those in the wild type B104 (P＜0.05), which was consistent with the late flowering phenotype of the mutants. 【Conclusion】CRISPR/Cas9 technology can be used to edit ZmFKF1 to obtain gene editing mutants, and the flowering time of these mutants is significantly delayed compared to wild type.

The Grain Dehydration Characteristics of the Main Summer Maize Varieties in Huang-Huai-Hai Region

XU TianJun,LÜ TianFang,ZHAO JiuRan,WANG RongHuan,XING JinFeng,ZHANG Yong,CAI WanTao,LIU YueE,LIU XiuZhi,CHEN ChuanYong,WANG YuanDong,LIU ChunGe

Scientia Agricultura Sinica. 2021, 54(4): 708-719. doi:10.3864/j.issn.0578-1752.2021.04.004

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【Objective】Grain mechanical harvesting is the developing direction of modern maize production in China. The moisture content at physiological maturity (PM) and grain dehydration rate after PM are the key factors for realizing maize grain mechanical harvesting. The aim of this study was to clarify the differences and influencing factors for the dehydration characteristics of different maize varieties, so as to provide a theoretical instruction for the breeding and extending of grain mechanical harvesting varieties.【Method】Taking18 main maize varieties in Huang-Huai-Hai region as research materials, the field experiment was conducted in 2017-2018, and the dynamics of maize grain moisture content were tested to study and clarify the differences and influencing factors for the dehydration characteristics.【Result】The moisture content of different maize varieties at PM and harvesting differed significantly, with an average of 30.67% (CV=2.58%) and 23.66% (CV=9.10%). There were significant differences between grain dehydration rate before and after PM of the tested varieties, with an average of 0.69%·d^-1 and 0.48%·d^-1, respectively. The average physical dehydration rate after PM of middle early maturing varieties was 0.55%·d^-1, which was 14.58% and 44.74% higher than that of middle maturing and middle late maturing varieties, respectively. The average yield of the tested varieties was 10 205.90 kg·hm^-2, with the range of 8 809.13-11 053.73 kg·hm^-2. The yield of middle maturity variety (10 484.25 kg·hm^-2) > middle late maturing variety (10 096.08 kg·hm^-2) > middle early maturing variety (9 522.81 kg·hm^-2), and Jingnongke728 and NK815 had the highest yield of 10 569.00 and 11 053.50 kg·hm^-2, respectively. Correlation analysis showed that the dehydration rate of grain was significantly positively correlated with the dehydration rate of leaves, bracts, rachis, stalk, whole plant and stem and wind speed. There was significantly positively correlated with atmospheric temperature and negatively correlated with atmospheric humidity. The varieties were divided into 4 types according to the grain dehydrate rate and yield by two-way average method. JNK728, JNK729, MC812 and MC121 were selected by using maize core inbred Jing2416 and its improved inbred Jing2418, which were characterized by early maturity and fast dehydrating rate as the male parent belongs to the fast dehydration rate and high yield type (The average growth period was 108.9 d; The average grain dehydration rate after physiological maturity was 0.57 %·d^-1; The moisture content of grain at harvest was 21.81%; The average yield was 10 811.33 kg·hm^-2).【Conclusion】According to the growth period, grain dehydration rat and yield level, the maize varieties of JNK728, MC812, JNK729 and MC121 were characterized by medium-early and medium maturity, fast dehydration rate and high yield, and could realize lower grain moisture content and higher yield level in Huang-Huai-Hai region.

The " Short, Dense and Early" Cultivation of Cotton in Xinjiang: History, Current Situation and Prospect

LOU ShanWei,DONG HeZhong,TIAN XiaoLi,TIAN LiWen

Scientia Agricultura Sinica. 2021, 54(4): 720-732. doi:10.3864/j.issn.0578-1752.2021.04.005

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Through years of research and practice, Xinjiang has formed a "short, dense, and early" cotton cultivation mode based on reasonable dense planting, dwarf plants, and early emergence and early maturity around 1994. The purpose of this mode was to promote early growth, fully utilize the technical methods of film mulching, drip irrigation, chemical regulation, mechanization, and informatization, and combine with cotton varieties, through dense planting and dwarf plants to promote the growth and development of cotton with synchronization of radiation use efficiency, and coordinate the contradiction between individuals and population of cotton. The group effect was used to increase the source and expand the sink to improve the effective photosynthetic area and photosynthetic production capacity, and to overcome adverse environmental impacts, such as low temperature in spring and rapid cooling in autumn. The "short, dense and early" cultivation mode could achieve early cotton maturity, high yield and high quality, which has increased Xinjiang's lint yield per hectare from 511.5 kg in 1981 to 1 200 kg in 1994. Under this mode, a high yield of 1 966.5 kg was achieved in 2019, and the average annual yield increased by more than 7.2%, which provided a mode for China and the world to explore high-yield cultivation modes. This article reviewed the formation and evolution of the "short, dense, early" cultivation mode, focusing on the content and high-yield mechanism of the mode, and prospects for the development of the mode, in order to provide a reference for the continuous innovation of cotton cultivation modes in the future.

The Expression Pattern and Interaction Analysis of the Homologues of Splicing Factor SC35 in Setosphaeria turcica

LI TianCong,ZHU Hang,WEI Ning,LONG Feng,WU JianYing,ZHANG Yan,DONG JinGao,SHEN Shen,HAO ZhiMin

Scientia Agricultura Sinica. 2021, 54(4): 733-743. doi:10.3864/j.issn.0578-1752.2021.04.006

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【Objective】The objective of this study is to obtain the homologous genes of the splicing factor SC35 in Setosphaeria turcica, and to analyze the interaction among them and the expression profiles during the different growth and development stages and infection process of the pathogen. It would lay the foundation for illustrating the relationship between the SC35 family members and fungal pathogenicity.【Method】Based on amino acid sequences of SC35 protein in Arabidopsis thaliana as probe sequences, online Blastp alignment was carried out in the S. turcica genome database to obtain candidate SC35 homologues. Then they were analyzed for conserved domain and phylogenetic relationship through bioinformatics procedures. The materials of S. turcica were collected at different infection stages on maize leaves and multiple developmental stages, such as hyphae, conidia, germ tubes, appressorium and penetration hyphae, to analyze the transcription levels of SC35 homologues through real-time quantitative PCR (qRT-PCR). And their interactions were verified in vitro by the yeast two-hybrid test.【Result】Eight SC35 genes of S. turcica were obtained, named as StSC1, StSC2, StSC3, StSC4, StSC5, StSC6, StSC7, and StSC8, respectively. All of them owned typical SR protein domains, besides StSC1 showed two RRM domains. They located at different physical locations of the genome and had no linkage. Phylogenetic analysis showed that the eight alternative splicing factors were distributed in different clades, with low homology. In the process of infection on maize leaves, gene StSC1, StSC2, StSC3, StSC4, StSC5, StSC6 and StSC8 were up-regulated, while StSC7 was down-regulated at 18 h after inoculation. StSC4 showed high expression activity at 6-18 h. In different development periods, the expression levels of StSC1 were extremely significant up-regulated (P＜0.001) in the appressorium and the formation of penetration hyphae, which were 24.44 and 8.25 times higher than those in the conidial period, but the others were down-regulated during the development. The yeast two-hybrid proved that StSC4 interacted with StSC6, StSC3 with StSC8, StSC3 with StSC4, and StSC8 with StSC4. 【Conclusion】The expression patterns of SC35 are different in the infection process and multiple developmental stages of S. turcica. StSC4 is actively expressed throughout the infection process. StSC1, StSC4 and StSC6 play important regulatory roles during the formation of appressorium and penetration hyphae. StSC4 and StSC6, StSC3 and StSC8, StSC3 and StSC4, StSC8 and StSC4 interact to regulate the formation of splicing complexes.

Expression in vitro of Metarhizium anisopliae Adhesin MAD1 and Its Effect on Inducing Response in Peanut

YAN DuoZi,CAI Ni,WANG Feng,NONG XiangQun,WANG GuangJun,TU XiongBing,ZHANG ZeHua

Scientia Agricultura Sinica. 2021, 54(4): 744-753. doi:10.3864/j.issn.0578-1752.2021.04.007

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【Objective】The fungus Metarhizium anisopliae has the characteristics of insect pathogenicity and plant symbiosis, and its adhesin MAD1 plays an important role in the infection to host insects. The objective of this study is to express MAD1 protein in eukaryotes in vitro, and to clarify the role of MAD1 in symbiosis between M. anisopliae and plants. 【Method】Unigenes of M. anisopliae transcriptome were used as the reference sequence to search and compare the homology in GenBank and design primers. The gene mad1 was cloned from the cDNA template of M. anisopliae IPPM010202. MAD1 protein sequence was translated by DNAMAN software. The online software ProtParam was used to predict the MAD1 protein composition of amino acid and its physicochemical characteristics. The protein structure was analyzed using SMART online program. The recombinant was constructed from eukaryotic expression vector with mad1 and Pichia pastoris transformation. The MAD1 protein was successfully expressed by methanol induction and the purified MAD1 was obtained by Ni-NTA affinity chromatography. The peanut roots were treated with immersion in the purified MAD1 solution (20 μg·mL^-1) for 0.5, 6, 12 and 24 h, and then the transcription levels of membrane receptor genes (CERK1, RPK), immune-related cascade related genes (MAPK, MMK1, CDPK) and transcription factor gene (MYB86), cell wall integrator gene (SCW1) as well as defense related genes (PTi1, RML1A) were detected by real-time fluorescence quantitative PCR (qPCR). 【Result】 The adhesive gene mad1 of M. anisopliae was cloned, with a total length of 2 136 bp, encoding 711 amino acids in molecular weight 74.8 kD. Bioinformatics analysis showed that the N-terminal of MAD1 has signal peptide, the C-terminal has a glycosylphosphatidyl inositol (GPI) anchor and contains CFEM functional domain and belongs to hydrophilic protein. The eukaryotic expression system with mad1 was successfully constructed and the active MAD1 protein was efficiently induced, expressed and purified. The results based on qPCR of peanut roots treated with purified MAD1 showed that, for 0.5 h, the root tip cells perceived the protein and activated the expression of membrane recognition receptor gene CERK1. In 0.5-6 h, the transcriptional levels of CERK1 and RPK were up-regulated, the transcription level of membrane-integrated gene SCW1 changed from down-regulation to up-regulation, while the transcriptional levels of MAPK, MMK1, CDPK and MYB86 were temporarily inhibited, and the transcriptional levels of defense genes PTi1 and RML1A were down-regulated, the root immune defense response was inhibited. In 6-12 h, the membrane recognition receptors maintained up-regulation, while the defensive gene RML1A reversed from down-regulation to strong up-regulation. In 12-24 h, CERK1 and RPK, PTi1 and RML1A were all up-regulated, MAPK, MMK1, CDPK and MYB86 appeared slightly up-regulated, the roots initiated immune defense response.【Conclusion】In the early stage of the interaction between M. anisopliae and peanut roots, the adhesin MAD1 protein activates the membrane receptor genes CERK1 and RPK recognition response in peanut at 6 h, while inhibits the expression of peanut immune cascade genes MAPK, CDPK, MMK1 and defense related genes such as PTi1 and RML1A, which is helpful for the colonization of M. anisopliae in peanut root tissue. Subsequently, it induces the up-regulation of expression of cell wall integrin gene SCW1, which participates in the repair and reconstruction of damaged cell wall on the root, and promotes the establishment of symbiotic relationship between M. anisopliae and peanut. Induction of plant immunosuppression and integral cell wall reconstruction may be important steps in the establishment of symbiotic relationship between M. anisopliae and plants.

Performance Study of Prothioconazole Microcapsules Prepared by Solvent Evaporation Method

CHEN Ge,CAO LiDong,XU ChunLi,ZHAO PengYue,CAO Chong,LI FengMin,HUANG QiLiang

Scientia Agricultura Sinica. 2021, 54(4): 754-767. doi:10.3864/j.issn.0578-1752.2021.04.008

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【Objective】The biodegradable material poly (3-hydroxybutyrate-co-4-hydroxybutyrate) (P (3HB-co-4HB)) was used as the wall material to prepare prothioconazole microcapsules. The effect of preparation process on the microcapsule size, pesticide loading and encapsulation efficiency was optimized. The microcapsules with good dispersion, small particle size, and high pesticide loading were screened out, and the preliminary researches on the release kinetics, photodegradation, and indoor biological activity on Sclerotium rolfsii were carried out. The purpose of this study is to provide a theoretical basis and technical support for improving the stability and utilization efficiency of prothioconazole in the environment.【Method】The solvent evaporation method was used to prepare prothioconazole microcapsules, and the effects of the mass ratio of core to wall material, volume ratio of oil to water, mass fraction of emulsifier and shearing speed on the particle size, pesticide loading and encapsulation efficiency of the microcapsules were investigated through a single factor test. Taking pesticide loading and particle size as the key technical indicators, the optimal preparation parameters were screened out through the L₉(3⁴) orthogonal test, which was further verified. The morphological and structural features, release performance and photodegradability of the microcapsules were determined by scanning electron microscope (SEM), fourier transform infrared (FTIR) spectrometer, and high performance liquid chromatography (HPLC). The toxicity of prothioconazole microcapsules on S. rolfsii was investigated by indoor bioassay.【Result】The mass ratio of core to wall material had a significant effect on the pesticide loading capacity of the microcapsules. As the ratio of core material increased, the loading capacity gradually increased. The volume ratio of oil to water, PVA mass fraction, and shearing speed had significant effects on the microcapsule particle size. As the shearing speed and PVA mass fraction increased, the microcapsule particle size gradually decreased. The volume ratio of oil to water had a great influence on the morphology and dispersion of microcapsules, and the influence of various factors on the encapsulation efficiency of the microcapsules was not significant. The optimal preparation parameters obtained through the L₉(3⁴) orthogonal test was as follows: the mass ratio of core to wall material of 1﹕5, volume ratio of oil to water of 1﹕5, PVA mass fraction of 2%, and shearing speed of 12 000 r/min. Under the optimal preparation process, spherical prothioconazole microcapsules with a particle size (D₅₀) of 3.32 μm and a span of 2.82 were prepared with a loading content of 15.52% and an encapsulation efficiency of 80.24%. Compared with prothioconazole technical material, the microcapsules had better sustained-release performance, and the release kinetics conformed to Fick’s diffusion law, presenting two processes of “burst release” followed by “sustained release”. The photostability of prothioconazole in the microcapsules in aqueous solution was enhanced, and the half-life of photolysis was doubled. The mycelial growth rate inhibition result showed that the fungicidal activity of prothioconazole microcapsules against S. rolfsii was equivalent to that of prothioconazole technical material.【Conclusion】Prothioconazole microcapsules with biodegradable material P (3HB-co-4HB) as a carrier were prepared, and different preparation processes affect the pesticide loading, dispersion state and particle size of microcapsules. The slow and sustained release and photostability are of great significance for reducing the amount of pesticide applied and improving the utilization efficiency of pesticide, which has potential application in control of peanut southern blight.

Analysis of Impacts and Regulation Differences on Soil N₂O Emissions from Two Typical Crop Systems Under Drip Irrigation and Fertilization

LEI HaoJie,LI GuiChun,KE HuaDong,WEI Lai,DING WuHan,XU Chi,LI Hu

Scientia Agricultura Sinica. 2021, 54(4): 768-779. doi:10.3864/j.issn.0578-1752.2021.04.009

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【Objective】The aim of this study was to ascertain the impact differences and emission reduction contribution of drip irrigation and fertilization on N₂O emissions from farmland of typical planting types in North China, and to clarify its comprehensive control mechanism, so as to provide scientific support and technical reserves for the optimization and control of regional agricultural production carbon and nitrogen and the promotion and application of drip fertilization technology in North China.【Method】Two typical crop planting modes (winter wheat-summer corn rotation and facility vegetable fields) were selected as the research objects, and four treatments were set, namely control (CK), conventional flood irrigation fertilization (FP), drip irrigation fertilization (FPD), and drip irrigation Optimized fertilization (OPTD). Continuous static observation and analysis of soil N₂O emissions from these two systems were performed by using automatic static chamber-gas chromatography.【Result】The changes of N₂O fluxes from the two crop systems were significantly positively correlated with the soil temperature of 5cm (P＜0.05), and both crop systems had the highest N₂O emission peak during the basal fertilizer period. In the greenhouse vegetable and food crop systems, the total cumulative N₂O emissions of FP treatment were the highest, reaching (5.47±0.23) and (1.70±0.02) kg N·hm^-2, respectively. For the N₂O emission intensity per unit yield, the FP treatment in the facility vegetable system was (159.72±2.47) g N·t^-1, which was much lower than the grain crop system (258.41±6.35) g N·t^-1. The focus of future N₂O emission reduction was still food crop production. Drip irrigation and fertilization could significantly reduce the total N₂O emissions of the two systems. Compared with FP treatment, drip irrigation and fertilization in facility vegetable systems could significantly reduce the total N₂O emissions by 19.0% (P＜0.05), while in food crop systems could be reduced by 35.0% (P＜0.05). In addition, when the nitrogen application rates of the two systems were reduced by 50% and 30%, the emission reduction contribution was expanded to 30.2% and 45.8%, respectively, while ensuring crop yields.【Conclusion】There were obvious differences in the characteristics of soil N₂O emissions from facility vegetable and food crop systems. The N₂O emission intensity of food crop production was significantly higher than that of facility vegetable production, and the further attention should be paid. At the same time, drip irrigation and fertilization technology could reduce N₂O emissions in two typical crop systems in North China farmland, but it had a greater contribution to N₂O emission reduction in the winter wheat-summer corn rotation system, and it had the potential for further application in the North China Plain.

Effects of Lime on Cadmium Accumulation of Double-Season Rice in Paddy Fields with Different Cadmium Pollution Degrees

ZHOU Liang,XIAO Feng,XIAO Huan,ZHANG YuSheng,AO HeJun

Scientia Agricultura Sinica. 2021, 54(4): 780-791. doi:10.3864/j.issn.0578-1752.2021.04.010

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【Objective】The purpose of this study was to explore the effects of applicating liming in the double-season rice in paddy fields with varying degrees of cadmium pollution, and then to provide a reference for rice safety production in different polluted paddy fields with early and late rice seasons.【Method】43 typical cadmium polluted paddy fields in counties (districts) of Hunan province were used as experimental sites, and the lime was applied according to the standard of 1 200 kg·hm^-2, to explore the change of four values such as soil pH, soil available Cd content, rice Cd content and rice enrichment coefficient, that were respectively researched in lightly polluted paddy fields (soil available Cd≤0.2 mg·kg^-1), moderately polluted paddy fields (0.2 mg·kg^-1soil available Cd≤0.4 mg·kg^-1), severely polluted paddy fields (0.4 mg·kg^-1soil available Cd≤0.6 mg·kg^-1), and super severely polluted paddy fields (soil available Cd>0.6 mg·kg^-1). The conventional cultivation was used as control.【Result】The multi-point experiment results showed that: (1) From the perspective of integral Cd polluted paddy fields, compared with conventional cultivation, the average Cd content in early and late rice was significantly reduced by applying lime, with a decrease of 31.0% and 28.6%, respectively. (2) From the perspective of different rice seasons in diverse pollution levels rice fields, the mean rice Cd content in paddy fields with moderately, severely, and seriously polluted rice fields in early rice seasons was decreased by 37.0%, 38.7% (P＜0.05) and 22.6%, respectively, compared with that in conventional cultivation. Application of lime could also reduce the mean Cd content of rice in lightly, moderately, severely and super severely polluted paddy fields in late rice season by 2.0% ,31.3% (P＜0.05), 31.8% and 22.9%, respectively. The lime could regulate Cd content in rice, because it could lower the Cd enrichment coefficient of rice in paddy fields with different degrees of pollution and the available Cd content in soil, ,and increase the soil pH. 【Conclusion】The application of lime could effectively control the mean Cd content of rice in the paddy fields polluted by light, moderate and severe Cd in early rice season and lightly Cd polluted paddy fields in late rice season to below the limit standard (0.2 mg·kg^-1). Therefore, on the basis of lime application and in combination with other measures to reduce Cd as the main ideas of “separating the degree of pollution in rice season” and “ late rice is prior to early rice” could improve the stability of controlling rice Cd content in field production and the economy of governance costs.

Correlation Analysis of Volatile Flavor Components and Metabolites Among Potato Varieties

LI KaiFeng,YIN YuHe,WANG Qiong,LIN TuanRong,GUO HuaChun

Scientia Agricultura Sinica. 2021, 54(4): 792-803. doi:10.3864/j.issn.0578-1752.2021.04.011

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【Objective】 This paper was aimed to investigate the metabolic compounds related to the change rules of volatile flavour components and to find out the potential metabolic precursor of the volatile flavour components in potato, so as to providing references to the flavour and quality improvement of potatoes. 【Method】 The gas chromatography-mass spectrometry (GC-MS) was used for the non-target detection of the tuber metabolome and post-curing volatile flavour components in the potato varieties (lines), including as Qingshu No.9, Huasong No. 7, Xisen No. 6, Houqihong and Jizhangshu No.12, D545, and D7277.【Result】 There were 22 significantly different volatile flavour compounds between the varieties among 7 main cultivation varieties of potatoes, most of which were found with high content in Jizhangshu No.12, and relatively low content in Huasong No.7. The clustering analysis with the compounds detected could classify the 7 varieties into 2 groups with distinct intensity of typical flavour; in group 1, the fragrance of Huasong No.7 after the curing method of Hot-air fry might be denser; in group 2, the baking flavour of potatoes of the variety of Jizhangshu No.12 was the most obvious. 69 metabolic compoundswere detected through the non-target metabolism detection of the 7 varieties of tubers, including 19 compounds differing significantly between the 2 variety groups. The main metabolite was related to the volatile flavour compounds to a certain extent. Methionine, alanine, threonine and sucrose were significantly and positively correlated with the methional and 2-methylfuran; while the proline and isoserine were significantly and negatively correlated with the E, Z-2, 6-nonadienaldehyde and E, E-2, 4-decadienal. 【Conclusion】 The content change of volatile flavour compounds was affected by the varieties, and methionine, alanine, threonine and sucrose were the metabolic compounds that were most likely to influence the constitution of the typical flavour of potatoes. The correlation analysis results could provide references for seeking the potential metabolic pathways.

Enantiomeric Analysis of Free Amino Acids in Different Teas

ZHU Yin,ZHANG Yue,YAN Han,LÜ HaiPeng,LIN Zhi

Scientia Agricultura Sinica. 2021, 54(4): 804-819. doi:10.3864/j.issn.0578-1752.2021.04.012

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【Objective】 Free amino acids are the main chemical compositions in teas, and they are closely related to the taste quality of tea. In account of the existence of stereogenic centers, most amino acids contain two enantiomers (D and L configurations) with obviously different taste characteristics and biological activities. However, due to the limitation of detection technology, the studies of L-amino acids were focused in the previous work, and the D-amino acids have been rarely studied. Therefore, the study on the enantiomers of free amino acids in teas is very important for deepening the theoretical system of tea chemistry and is beneficial for improving and controlling of tea taste quality. 【Method】 In this study, the separation performance of free amino acid enantiomers (Chiral HPLC, Silanization of MTBSTFA and Esterification PFP acylation) was compared by using three different analytical methods. A total of 15 pairs of amino acids, including alanine, valine, threonine, isoleucine, leucine, proline, serine, cysteine, aspartic acid, methionine, phenylalanine, glutamic acid, tyrosine, lysine and tryptophan, were effectively separated by using esterification- pentafluoropropionic acylation combined with chiral gas chromatography-mass spectrometry (GC-MS). Furthermore, the efficient derivatization, qualitative and quantitative analysis approaches were established, and the distribution regularities of amino acids enantiomers in 11 representative commercial white, oolong and pu-erh teas were investigated. 【Result】 The derivatization conditions were showed as follows: the reaction mixture was esterified at 100oC for 105 min, and then further acylated by pentafluoropropionic anhydride using tetrahydrofuran as the solvent at 100oC for 10 min. Recoveries of the free amino acid enantiomers were ranged from 75.26% to 123.6% (low concentration) and from 81.23% to 121.8% (high concentration), and the corresponding RSDs were ranged from 2.09% to 13.12% (low concentration) and from 1.48% to 10.59% (high concentration). The analysis results indicated that 1-2 enantiomers of most amino acids could be detected, and D-amino acids were frequently distributed in most tea samples, especially D-threonine, D-aspartic acid, D-cysteine, D-phenylalanine, and the mixture of D-theanine, D-glutamic acid and D-glutamine. On the contrary, D-methionine, D-lysine and D-tryptophan were unable to be detected in all samples. As for the detailed content distributions, in addition to the mixture of L-theanine, L-glutamic acid and L-glutamine and L-aspartic acid, L-cysteine (1.48-2.08 mg?g^-1), D-cysteine (1.46-1.49 mg?g^-1) and D-aspartic acid (1.02-1.14 mg?g^-1) presented higher contents in white teas, L-cysteine (1.52-1.70 mg?g^-1), D-cysteine (1.45-1.49 mg?g^-1), L-serine (1.03-1.50 mg?g^-1), L-methionine (1.03-1.52 mg?g^-1), L-tyrosine (1.32-1.35 mg?g^-1) and D-aspartic acid (1.01-1.15 mg?g^-1) were abundant in oolong teas. In pu-erh teas, no significant differences on the contents between the mixture of L-theanine, L-glutamic acid and L-glutamine (1.04 mg?g^-1) and other amino acids, and L-threonine, L-tryptophan, and L-serine presented relatively high content levels ranging from 0.61 to 0.84 mg?g^-1. The content distribution of the total amino acids were presented the following tendency: white tea (up to 40.61 mg?g^-1) > oolong tea (up to 25.43 mg?g^-1) > pu-erh tea (8.01 mg?g ^-1). Moreover, multivariate statistical analysis results indicated that the mixture of L-theanine, L-glutamic acid and L-glutamine, L-tryptophan, L-aspartic acid, L-tyrosine and L-methionine showed significant content differences, and the first three enantiomers were abundant in white teas, and other compounds were rich in oolong teas.【Conclusion】 D-amino acids could be detected in most teas, indicating their potential impact on the taste quality of tea infusion. However, no significant differences on the content distribution of D-amino acids were observed among different kinds of teas used in our study.

Insight into the Impact of Heat Treatment on the Foamability and Structure of Gliadin Colloidal Particles

WANG LiFeng,ZHU Jie,XIONG WenFei,ZHAO Meng,YUAN Jian,JU XingRong

Scientia Agricultura Sinica. 2021, 54(4): 820-830. doi:10.3864/j.issn.0578-1752.2021.04.013

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【Objective】Gliadin derived from wheat gluten has strong surface hydrophobic properties, and the gliadin colloid particles prepared by ethanol-water solution anti-solvent exhibit outstanding foaming ability and stability. However, the foam properties of gliadin under the action of heat have not been revealed yet. Therefore, in order to further promote the application of gliadin particles in real food systems, the effects of different heating temperatures and heating times on the foaming ability and stability of gliadin particles were studied in this paper. 【Method】After treating the gliadin at different temperatures (50, 70 and 90℃) for 15, 30 and 60 minutes, the gliadin particles were prepared by the anti-solvent method, and the foaming ability and foam stability were measured. By measuring the size, zeta potential, protein solubility, atomic force microscopy, infrared spectroscopy, fluorescence spectroscopy, circular dichroism, ultraviolet spectroscopy, DSC and small-angle X-ray scattering of the heat-treated wheat gliadin particles, the changing law of its surface morphology and microstructure were analyzed. 【Result】The results showed that the foaming ability and foam stability of the heat-treated wheat gliadin particles increased by 25% and 85%, respectively. With the increase of the heating temperature and the extension of the heating time, the gliadin particles had partially aggregated; the particle size increases, and it mainly distributed around 105-122 nm, and the zeta potential decreases; the degree of aggregation became greater at 90℃. The heating temperature had no obvious effect on protein solubility, but the solubility of protein has been significantly improved with the increase of heating time; the thermal effect exposed the hydrophobic amino acids inside the protein molecule, resulting in an increase in surface hydrophobicity; the content of disulfide bonds decreases, and the content of free sulfhydryl groups had no significant difference. The reason might be that the SH/SS exchange reaction of the prolamin occurs during the heating process. High temperature treatment changed the secondary structure of wheat gliadin. The fluorescence intensity of the protein at 90℃ decreased, the β-sheet content decreased, while the irregular curl content increased, and the protein was highly stretched, accompanied by partial unfolding. DSC results showed that the highest energy of wheat gliadin particles decreased from 54.33 mW to about 3 mW, the spectrum after heating was relatively flat, and the protein conformation tended to be amorphous with the extension of heating time.【Conclusion】The thermal effect caused the wheat gliadin particles to aggregate, and the exposure of the hydrophobic groups in the protein enhanced the hydrophobicity of the particle surface. Heat treatment improved the structural flexibility of the wheat gliadin (especially the 90℃ treatment), which was more conducive to the formation of stability. The interfacial film could better stabilize the foam and effectively improve the foam characteristics of the wheat gliadin colloid particles, which had outstanding practical significance for enhancing its application in the food industry.

Effects of Whole Plant Corn Silage Ratio in Diet on Growth Performance, Rumen Fermentation, Nutrient Digestibility and Serological Parameters of Dorper×Hu Crossbred Female Lambs

WANG JinFei,YANG GuoYi,FAN ZiHan,LIU Qi,ZHANG PengCheng,REN YouShe,YANG ChunHe,ZHANG ChunXiang

Scientia Agricultura Sinica. 2021, 54(4): 831-844. doi:10.3864/j.issn.0578-1752.2021.04.014

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【Objective】The aim of this study was to explore the effects of different proportions of the whole-plant corn silage on growth performance, rumen fermentation, nutrient digestibility, serum biochemical indexes, antioxidative enzymes’ activities, and immune function at different stages of Doper ×Hu crossbred female lambs.【Method】Seventy-two female lambs (Dorper sheep×Hu sheep) with good body condition and an approximate weight (16 ± 1.5) kg were selected and randomly divided into four groups, each group with six replicates, each replicate with three lambs. Group I as the control group was fed with peanut vine as the basis of roughage source in diet, Group Ⅱ, Ⅲ, and Ⅳ for experimental groups were respectively fed 20%, 40%, and 60% the whole-plant corn silage (dry matter basis) instead of peanut vine in the diet. The experiment lasted for 115 days, of which the pre-test period was 15 days and the formal test period was 100 days, including 90 days for a feeding trial and 10 days for a digestibility trial.【Result】(1) Compared with I group, average daily gain in group III increased significantly (P＜0.05) from 1 to 30 days, and feed conversion ratio significantly reduced (P＜0.05) from 1 to 30 days and 1 to 90 days. (2) The addition of whole-plant corn silage in the diet could improve the rumen fermentation of lambs. With the increase of proportions of whole-plant corn silage, acetate, butyrate, and acetate/propionate decreased significantly in rumen fluid of lambs (P＜0.05); the ratio of propionate increased significantly (P＜0.05); group IV had a considerably higher concentration of NH₃-N than group I, II, and III in rumen fluid of lambs (P＜0.05). (3) The apparent digestibility of DM, GE of group III and IV was significantly higher than those of group I (P＜0.05), the apparent digestibility of OM, the nitrogen of group II, III, and IV were markedly higher than those of group I (P＜0.05). With the increase of proportions of whole-plant corn silage in diet, the fecal nitrogen excretion of lambs decreased significantly (P＜0.05). And the fecal nitrogen excretion of group II, III, and IV was markedly lower than that of group I (P＜0.05). Lambs in group IV had significantly higher urinary nitrogen excretion (P＜0.05), which resulted in a lower nitrogen retention rate than that of group III (P＜0.05). (4) Serum glucose concentration in group III was significantly higher than in group I and II on the 90th day (P＜0.05). (5) Group III had a considerably higher serum T- AOC and lower serum MDA concentration than group I did on the 90th day (P＜0.05); serum SOD and GSH-Px activities in group IV were significantly higher than those in group I on the 60th day and 90th day(P＜0.05). Group IV had significantly higher serum IgA and IgM concentration (P＜0.05) and significantly lowered the levels of TNF-α than group I did on the 60th day and 90th day (P＜0.05).【Conclusion】Forty percentage of whole-plant corn silage in diet could improve the rumen fermentation, increase nutrient digestibility, enhance the antioxidant ability and immunity, and promote the healthy growth of lambs.

Expression Patterns of TETs and Programmed Cell Death Related Genes in Oviduct and Uterus of Early Pregnancy Goats

ZHAO Le,YANG HaiLi,LI JiaLu,YANG YongHeng,ZHANG Rong,CHENG WenQiang,CHENG Lei,ZHAO YongJu

Scientia Agricultura Sinica. 2021, 54(4): 845-854. doi:10.3864/j.issn.0578-1752.2021.04.015

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【Objective】The ten-eleven translocation (TET) protein family play crucial roles in embryo and placental development. The balance of cell death between oviduct and endometrial would affect the development of gametes and early embryos. In this study, the expression patterns and their potential regulatory role of TETs, programmed cell death related genes and blastocyst implantation-related genes in oviduct and uterus of goats in early pregnancy were mainly explored, so as to provide a basis for studying development of gametes and early embryos.【Method】In this study, the uterus and oviduct of Hechuan white goats during non-pregnant 19 days (C19) and pregnant 19 days (P19) were collected, and then the morphology uterine horn was showed by HE staining. Then, the genes, including blastocyst implantation-related genes (Wnt5a, OPN, VEGFA), TET family genes (TET1, TET2, TET3), programmed cell death-related genes (BAX, BCL2, Caspase9, GSDMD, NLRP3 and Caspase1) and inflammation-related genes (NF-kB、TNF-α) were detected by qRT-PCR. Finally, the correlation between TETs and programmed cell death-related genes expression were analyzed.【Result】The results showed that the gland number increased, and its morphology varied in uterine horn in P19 group. Compared with C19 group, OPN gene expression and Bax/BCL2 in uterine horn tissue were significantly increased in P19 group (P＜0.01), however, there was no significant difference between Wnt5a and VEGFA. TET family genes were expressed in both oviduct and uterus. The expression of TET1 and TET2 was significantly higher than TET3 in uterine horn (P＜0.01). TET2 (P＜0.01), NF-kB (P＜0.01), Caspase1 (P＜0.01), and GSDMD (P＜0.05) gene were significantly higher in P19 group of oviduct tissue. Correlation analysis showed that the relationship between TET2 and TET3 (P＜0.01), NF-kB (P＜0.01) and Caspase9 (P＜0.01) in oviduct tissue were positive correlation. The TET3 were negative correlated with VEGFA (P＜0.05) and WNT5a (P＜0.01). Blastocyst implantation-related genes OPN was positively correlated with Caspase1 (P＜0.01), NLRP3 (P＜0.05) and GSDMD (P＜0.05) in uterine horn. The GSDMD was positively correlated with Bax (P＜0.05), Caspase1 (P＜0.01) and NLRP3 (P＜0.05) in oviduct and uterus horn.【Conclusion】It was indicated that there was correlation between TETs and programmed cell death related genes. So, it was speculated that TET family genes and programmed cell death related genes played a key role in regulating the development of oviduct, zygote, early embryo and uterus. The key genes for embryo implantation of OPN were significantly correlated with pyroptosis related genes, suggesting that proptosis had a certain influence on the attachment of early embryos. These finding provided heretical references for studying the effect of TETs family genes and programmed cell death related genes on oviduct and uterine horn of early pregnancy.

The Induction of Unfolded Protein Response in Tembusu Virus Infected Ducklings

ZHAO DongMin,HUANG XinMei,ZHANG LiJiao,LIU QingTao,YANG Jing,HAN KaiKai,LIU YuZhuo,LI Yin

Scientia Agricultura Sinica. 2021, 54(4): 855-863. doi:10.3864/j.issn.0578-1752.2021.04.016

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【Objective】 The aim of this study was to determine the signal pathways (PERK, IRE1 and ATF6) of unfolded protein response induced by tembusu virus in ducklings, so as to provide a theoretical basis for elucidating the pathogenesis of TMUV. [Method] 1-day-old SPF ducklings were infected intraperitoneally with TMUV (JS804). Ducklings inoculated in the same manner with equal volume of RMPI-1640 were used as negative control. Five ducklings from each group were euthanized at 12, 24, 36 h and 48 h post infection, and their brains, livers and hearts were collected. The total RNAs were extracted from collected tissues by using total RNA extraction kit. Then the total RNAs were reverse transcribed into cDNA. Specific target genes representing the three known pathways of unfolded protein response were selected, and the primers were designed based on the published GenBank sequence. The relative expression of target genes was quantified by real time PCR. Real time PCR data were analyzed using the comparative Ct method (2^-ΔΔCt). GAPDH was chosen as a reference gene for internal control. 【Result】 In three organs from infected ducklings, it was observed that the viral titers were the highest in the liver, lower in the heart, and the lowest in the brain. The unfolded protein response was characterized by upregulated expression of GRP78. The relative expression of GRP78 in the brain and liver were persistently upregulated and reached a peak at 36 h post infection (4.21 fold and 10.14 fold, respectively). GRP78 expression in the heart was upregulated transiently at 36 h post infection (1.32 fold). ATF4 expression represented the activation of PERK pathway. The ATF4 expression in the liver and brain were persistently upregulated from 24 h and 36 h post infection to 48 h post infection respectively, and peaked at 36 h post infection (2.71 fold and 6.02 fold, respectively). However, the upregulation of ATF4 expression in the heart was observed at 36 h post infection (1.57 fold). The activation of IRE pathway was characterized by XBP1s. In the liver, the expression level of XBP1s increased most significantly (9 fold). In the brain, the expression level of EDEM enhanced most significantly (3.87 fold) and persistently upregulated from 12 to 48 h post infection. Comparing to negative control, the expression of ATF6 pathway marker GRP94 and XBP1u were upregulated in three tissues, which reached a peaked at 36 h post infection, although expression profiles of GRP94 and XBP1u were different at indicated time points. 【Conclusion】 It was the first report that TMUV infection induced three branches of unfolded protein response in ducklings, and these results might be helpful for understanding the interaction between tembusu virus infection and host response.

Construction and Annotation of Ascosphaera apis Full-Length Transcriptome Utilizing Nanopore Third-Generation Long-Read Sequencing Technology

DU Yu,ZHU ZhiWei,WANG Jie,WANG XiuNa,JIANG HaiBin,FAN YuanChan,FAN XiaoXue,CHEN HuaZhi,LONG Qi,CAI ZongBing,XIONG CuiLing,ZHENG YanZhen,FU ZhongMin,CHEN DaFu,GUO Rui

Scientia Agricultura Sinica. 2021, 54(4): 864-876. doi:10.3864/j.issn.0578-1752.2021.04.017

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【Objective】Purified mycelia sample (Aam) and spore sample (Aas) were sequenced using third-generation nanopore long-read sequencing technology, followed by construction and annotation of high-quality full-length transcriptome.【Method】Aam and Aas were respectively sequenced using Oxford Nanopore PromethION platform. Guppy software was used to conduct base calling of raw reads. Clean reads were obtained after filtering out short fragments and low-quality raw reads. Full-length transcripts were identified by recognizing primers at both ends of clean reads. Full-length transcripts were aligned to Nr, Swissprot, KOG, eggNOG, Pfam, GO and KEGG databases to gain corresponding annotations. Four approaches such as CPC, CNCI, CPAT, and Pfam were used to predict lncRNAs, and the intersection was deemed to be high-reliability lncRNAs.【Result】In total, 6 321 704 and 6 259 727 raw reads were yielded from nanopore sequencing of Aam and Aas, and after quality control, 5 669 436 and 6 233 159 clean reads were obtained, including 4 497 102 (79.32%) and 4 963 101 (79.62%) full-length clean reads. Additionally, 9 859 and 16 795 non-redundant full-length transcripts were identified, with a N50 of 1 482 and 1 658 bp, an average length of 1 187 and 1 303 bp, and a maximum length of 6 472 and 6 815 bp, respectively. Venn analysis showed that 6 512 non-redundant full-length transcripts were shared by Aam and Aas, while 3 347 and 10 283 ones were specific for Aam and Aas, respectively. Besides, a total of 20 142 full-length transcripts were identified in Aam and Aas, among them 20 809, 11 151, 17 723, 12 164, 11 340 and 9 833 full-length transcripts could be annotated to Nr, KOG, eggNOG, Pfam, GO and KEGG databases, respectively. Most of full-length transcripts were annotated to A. apis, Polytolypa hystricis and Histoplasma capsulatum. Moreover, GO database annotation demonstrated that the above-mentioned full-length transcripts could be annotated to 45 functional terms, involving in cell component-associated terms such as cell part, cell and organelle; molecular function-associated terms such as catalytic activity, binding and transporter activity; and biological process-associated terms such as cellular processes, metabolic processes and single-organism processes. KEGG database annotation indicated that these full-length transcripts could be annotated to 49 pathways, including biosynthesis of antibiotics, ribosome, biosynthesis of amino acid, carbon metabolism, spliceosome and so on. In addition, 648 lncRNAs were identified, including 480 long intergenic RNAs (lincRNAs), 119 anti-sense lncRNAs and 49 sense lncRNAs. 【Conclusion】The first high-quality full-length transcriptome was constructed and annotated in this work, which offers a key basis for exploration of the complexity of A. apis transcriptome, improvement of sequence and functional annotation of reference genome and further study on isoforms’ function of A. apis.

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