Rv3435c重组耻垢分枝杆菌感染小鼠RAW264.7巨噬细胞的转录组分析

doi:10.11843/j.issn.0366-6964.2025.09.043

畜牧兽医学报 ›› 2025, Vol. 56 ›› Issue (9): 4657-4672.doi: 10.11843/j.issn.0366-6964.2025.09.043

Rv3435c重组耻垢分枝杆菌感染小鼠RAW264.7巨噬细胞的转录组分析

刘昕玥¹, 李丹妮¹, 宗颖^2,3,4, 时坤^2,3,4, 李健明^2,3,4, 刁乃超^2,3,4, 曾范利^2,3,4,5*, 杜锐^2,3,4,5*

1. 吉林农业大学动物医学院/动物科技学院, 长春 130118;
2. 吉林农业大学中药材学院, 长春 130118;
3. 梅花鹿药用资源利用关键技术研究室, 长春 130118;
4. 吉林省梅花鹿高效养殖和产品开发技术工程研究中心, 长春 130118;
5. 教育部动物生产及产品质量安全重点实验室, 长春 130118

收稿日期:2024-12-02 发布日期:2025-09-30
通讯作者: 曾范利，主要从事经济动物疫病防治及人兽共患病防治研究，E-mail:zengfanli@jlau.edu.cn；杜锐，主要从事经济动物疫病防治及人兽共患病防治研究，E-mail:durui@jlau.edu.cn
作者简介:刘昕玥(1999-)，女，山东莱州人，硕士生，主要从事经济动物疫病防治研究，E-mail:2892144827@qq.com
基金资助:
国家自然科学基金联合基金项目(U23A20237)

Transcriptome Analysis of RAW264.7 Macrophages Infected with Rv3435c Recombinant Mycobacterium smegmatis

LIU Xinyue¹, LI Danni¹, ZONG Ying^2,3,4, SHI Kun^2,3,4, LI Jianming^2,3,4, DIAO Naichao^2,3,4, ZENG Fanli^2,3,4,5*, DU Rui^2,3,4,5*

1. College of Veterinary Medicine/College of Animal Science and Technology, Jilin Agricultural University, Changchun 130118, China;
2. College of Chinese Medicine Materials, Jilin Agricultural University, Changchun 130118, China;
3. Research Laboratory on Key Technology of Development and Resource Utilization Sika Deer, Changchun 130118, China;
4. Jilin Province Sika Deer Efficient Breeding and Product Development Technology Engineering Research Center, Changchun 130118, China;
5. The Ministry of Education Key Laboratory of Animal Production and the Product Quality and Safety, Changchun 130118, China

Received:2024-12-02 Published:2025-09-30

摘要/Abstract

摘要： 前期研究发现，异源表达Rv3435c基因的耻垢分枝杆菌显著抑制巨噬细胞的炎症表达，为了探究Rv3435c基因在该过程行使的功能，使用二代测序分析Rv3435c重组耻垢分枝杆菌感染RAW264.7细胞的转录组差异。结果显示：通过RT-qPCR检测6、12、24、48 h 的IL-1β、TNF-α、IL-6 mRNA表达水平，选择表达量最高的12 h作为转录组测序时间点。将测序获得的原始数据进行数据质量分析及比对结果统计，确定数据质量良好，进行样本关系分析，提示Rv3435c显著改变了小鼠巨噬细胞的基因表达情况。将数据过滤比对共得到14 077个差异基因，其中，有278个显著上调表达基因和118个显著下调表达基因。对差异基因进行GO注释，得到的差异基因主要富集在免疫系统过程、对外部刺激的反应、防御响应等方面。对差异基因进行KEGG通路富集分析，主要富集在环境信息处理、细胞过程、人类疾病、有机体系统几方面。通过蛋白互作分析筛选出核心基因10个，分别为IL-6、IL-1β、CCL2、Mmp9、Spp1、Itgam、Cdc20、Ccna2、Kif11、Ccnb2。对差异基因进行荧光定量PCR试验，验证转录组结果可信，根据筛选出的DEGs和RT-qPCR结果，推测SPP1是Rv3435c行使功能的潜在靶点，并通过RT-qPCR、ELISA、Western blot验证结果。本文为结核分枝杆菌Rv3435c基因的功能探索提供了重要的数据支持。

关键词: 结核分枝杆菌, Rv3435c基因, 耻垢分枝杆菌, 基因注释, 转录组

Abstract: Previous studies have found that Mycobacterium smegmatis, which heterogeneically expresses the Rv3435c gene, can significantly inhibit the inflammatory expression of macrophages. In order to explore the function of Rv3435c gene in this process, second-generation sequencing was used to analyze the transcriptome differences of RAW264.7 cells infected by Rv3435c recombinant Mycobacterium smegmatis. Results were as follows: The mRNA expression levels of IL-1β, TNF-α and IL-6 at 6, 12, 24 and 48 h were detected by RT-qPCR, and 12 h with the highest expression levels was selected as the transcriptomic sequencing time point. The original data obtained by sequencing were analyzed for data quality and compared with the result statistics, and the data quality was confirmed to be good. The sample relationship analysis suggested that Rv3435c significantly changed the gene expression of mouse macrophages. A total of 14 077 differential genes were obtained by filtering and comparing the data, of which 278 were significantly up-regulated and 118 were significantly down-regulated. The differential genes obtained by GO annotation are mainly concentrated in immune system process, response to external stimulus, defense response and so on. KEGG pathway enrichment analysis was carried out for differential genes, mainly concentrated in environmental information processing, cellular processes, human diseases, and organic systems. Ten core genes were identified by protein interaction analysis, which were IL-6, IL-1β, CCL2, Mmp9, Spp1, Itgam, Cdc20, Ccna2, Kif11 and Ccnb2. Fluorescent quantitative PCR experiments were performed on the differential genes to verify the reliability of the transcriptome results. According to the selected DEGs and RT-qPCR results, we speculated that SPP1 was a potential target for the function of Rv3435c, and the results were verified by RT-qPCR, ELISA and Western blot. This study provides important data support for the function exploration of Rv3435c gene of Mycobacterium tuberculosis.

Key words: Mycobacterium tuberculosis, Rv3435c gene, Mycobacterium smegmatis, gene annotation, transcriptome

中图分类号:

S852.618

刘昕玥, 李丹妮, 宗颖, 时坤, 李健明, 刁乃超, 曾范利, 杜锐. Rv3435c重组耻垢分枝杆菌感染小鼠RAW264.7巨噬细胞的转录组分析[J]. 畜牧兽医学报, 2025, 56(9): 4657-4672.

LIU Xinyue, LI Danni, ZONG Ying, SHI Kun, LI Jianming, DIAO Naichao, ZENG Fanli, DU Rui. Transcriptome Analysis of RAW264.7 Macrophages Infected with Rv3435c Recombinant Mycobacterium smegmatis[J]. Acta Veterinaria et Zootechnica Sinica, 2025, 56(9): 4657-4672.

参考文献

[1] ANES E, PIRES D, MANDAL M, et al. ESAT-6 a Major virulence factor of mycobacterium tuberculosis[J]. Biomolecules, 2023, 13(6): 968.
[2] KOCH A, MIZRAHI V. Mycobacterium tuberculosis[J]. Trends Microbiol, 2018, 26(6): 555-556.
[3] KANABALAN R D, LEE L J, LEE T Y, et al. Human tuberculosis and Mycobacterium tuberculosis complex: A review on genetic diversity, pathogenesis and omics approaches in host biomarkers discovery[J]. Microbiol Res, 2021, 246: 126674.
[4] EHRT S, SCHNAPPINGER D, RHEE K Y. Metabolic principles of persistence and pathogenicity in Mycobacterium tuberculosis [J]. Nat Rev Microbiol, 2018, 16(8): 496-507.
[5] World Organisation for Animal Health. Report of the meeting of the ad hoc group on alternative strategies for the control and elimination of Mycobacterium tuberculosis complex infection (MTBC) in livestock[C/OL]. Paris, France, 2024.[2025-07-02]https://www.woah.org/app/uploads/2024/02/en-20240222-ahg-mtb-report.pdf.
[6] 世界动物卫生组织. 哺乳动物、禽、蜜蜂A和B类疾病诊断试验和疫苗标准手册[M].北京:中国农业出版社，2002:337-349.
World Organisation for Animal Health. Manual of diagnostic tests and vaccines for terrestrial animals (mammals, birds and bees): List A and B diseases[M]. Beijing: China Agriculture Press, 2002: 337-349. (in Chinese)
[7] ZHAO D, SONG Y H, LI D, et al. Mycobacterium tuberculosis Rv3435c regulates inflammatory cytokines and promotes the intracellular survival of recombinant Mycobacteria[J]. Acta tropica, 2023, 246: 106974.
[8] SINGH A K, CARETTE X, POTLURI L P, et al. Investigating essential gene function in Mycobacterium tuberculosis using an efficient CRISPR interference system[J]. Nucl Acids Res, 2016, 44(18): e143.
[9] LU Q, ZHANG W, FANG J, et al. Mycobacterium tuberculosis Rv1096, facilitates mycobacterial survival by modulating the NF-κB/MAPK pathway as peptidoglycan N-deacetylase[J]. Mol Immunol, 2020, 127: 47-55.
[10] XIA A, LI X, QUAN J, et al. Mycobacterium tuberculosis Rv0927c inhibits NF-κB pathway by downregulating the phosphorylation level of IκBα and enhances Mycobacterial survival[J]. Front Immunol, 2021, 12: 721370.
[11] 韩梅, 韩璞, 邹静波. 结核分枝杆菌感染与细胞因子的关系[J]. 检验医学与临床，2021, 18(2): 270-272.
HAN M, HAN P, ZOU J B, et al. Relationship between Mycobacterium tuberculosis infection and cytokines[J]. Laboratory Medicine and Clinical, 2021, 18(2): 270-272. (in Chinese)
[12] 付加芳，张佩佩，古苑欣，等.结核分枝杆菌Rv1057基因对巨噬细胞感染早期细胞因子表达的影响分析[J].生命科学研究，2017，21(6):494-500.
FU J F, ZHANG P P, GU Y X, et al. Effect of Mycobacterium tuberculosis Rv1057 gene on the cytokine expression in the early stage of macrophage infection[J]. Life Science Research, 2017, 21(6): 494-500. (in Chinese)
[13] 张泽霖. Viperin通过IRAK1-TRAF6-TAK1负调节炎症细胞因子和NO促进结核分枝杆菌感染[D]. 广州：南方医科大学, 2020.
ZHANG Z L. Viperin impairs innate immune response through IRAK1-TRAF6-TAK1 axis to promote Mycobacterium tuberculosis infection[D]. Guangzhou: Southern Medical University, 2020. (in Chinese)
[14] 邓傲竹, 张少言, 冯雅, 等. 结核分枝杆菌与巨噬细胞：从感染机制到免疫逃逸策略的研究进展[C]//第35届中国防痨协会全国学术大会暨第四届中国防痨科技颁奖大会. 湖州, 2024.
DENG A Z, ZHANG S Y, FENG Y, et al. Mycobacterium tuberculosis and macrophages: Research progress from infection mechanisms to immune escape strategies[C]//Proceedings of the 35th National Academic Conference of China Anti-Tuberculosis Association and the 4th China Anti-Tuberculosis Science and Technology Award Conference. Huzhou, Zhejiang, China, 2024. (in Chinese)
[15] 李娜, 宋银娟, 储岳峰. 结核分枝杆菌免疫逃逸机制研究进展[J]. 科学通报，2024, 69(Z1): 531-41.
LI N, SONG Y J, CHU Y F. Research advances in immune evasion mechanisms of Mycobacterium tuberculosis[J]. Chinese Science Bulletin, 2024, 69(Z1): 531-541. (in Chinese)
[16] 李玉洁, 余海燕, 杨雨婷, 等. 结核分枝杆菌分泌蛋白早期分泌性抗原6(ESAT-6)的免疫学性质及其在新型疫苗中作用的研究进展[J]. 细胞与分子免疫学杂志，2024, 40(1): 89-94.
LI Y J, YU H Y, YANG Y T, et al. Immunological properties of Mycobacterium tuberculosis-secreted early secretory antigenic target 6 (ESAT-6) and its role in novel vaccines[J]. Cellular & Molecular Immunology, 2024, 40(1): 89-94. (in Chinese)
[17] AKASHI S, SUZUKAWA M, TAKEDA K, et al. IL-1RA in the supernatant of QuantiFERON-TB Gold In-Tube and QuantiFERON-TB Gold Plus is useful for discriminating active tuberculosis from latent infection[J]. J Infect Chemother, 2021, 27(4): 617-624.
[18] 马子淳, 尚媛媛, 逄宇,等. 趋化因子用于结核病诊断的研究进展[J]. 中国防痨杂志，2023, 45(3): 305-310.
MA Z C, SHANG Y Y, PANG Y, et al. Research progress on the application of chemokines in the diagnosis of tuberculosis[J]. Chinese Journal of Antituberculosis, 2023, 45(3): 305-310. (in Chinese)
[19] WANG T, QUIJADA D, AHMENDA T, et al. Targeting CCRL2 enhances therapeutic outcomes in a tuberculosis mouse model[J]. bioRxiv, 2024: 2024.09.23.614576.
[20] KUMAR N P, MOIDEEN K, NANCY A, et al. Plasma chemokines are baseline predictors of unfavorable treatment outcomes in pulmonary tuberculosis[J]. Clin Infect Dis, 2021, 73(9): e3419-e3427.
[21] HUANG H. Matrix metalloproteinase-9 (MMP-9) as a cancer biomarker and MMP-9 biosensors: recent advances[J]. Sensors (Basel), 2018, 18(10):3249.
[22] SEITZ-HOLLAND J, ALEMN-GÓMEZ Y, CHO K I K, et al. Matrix metalloproteinase 9 (MMP-9) activity, hippocampal extracellular free water, and cognitive deficits are associated with each other in early phase psychosis[J]. Neuropsychopharmacology, 2024, 49(7): 1140-1150.
[23] 李军霞, 赵青, 何红彦, 等. 基质金属蛋白酶-9与血脑屏障和结核性脑膜炎[J]. 中国感染与化疗杂志，2017, 17(4): 463-467.
LI J X, ZHAO Q, HE H Y, et al. Matrix metalloproteinase-9 and its role in the blood-brain barrier and tuberculous meningitis[J]. Chinese Journal of Infection and Chemotherapy, 2017, 17(4): 463-467. (in Chinese)
[24] 杨泽伟, 冯飞, 杨颖, 等. 脑脊液ESAT-6、ADA、INF-γ、MMP-9检测在结核性脑膜炎诊断及转归中的应用价值[J]. 山东医药，2018, 58(18): 56-58.
YANG Z W, FENG F, YANG Y, et al. Diagnostic and prognostic value of ESAT-6, ADA, INF-γ, and MMP-9 in cerebrospinal fluid for tuberculous meningitis[J]. Shandong Medical Journal, 2018, 58(18): 56-58. (in Chinese)
[25] 王霞, 黄健, 牛文一, 等. IFN-γ、MMP-9水平在肺结核中的表达及与其病情活动性的相关性[J]. 分子诊断与治疗杂志，2024, 16(7): 1372-1376.
WANG X, HUANG J, NIU W Y, et al. Expression of IFN-γ and MMP-9 in pulmonary tuberculosis and their correlation with disease activity[J]. Journal of Molecular Diagnostics and Therapy, 2024, 16(7): 1372-1376. (in Chinese)
[26] 韦爽. 血清MMP-9、MMP-2、TIMP-1及TIMP-2在婴幼儿肺炎中的表达水平及临床意义[D]. 遵义：遵义医科大学, 2023.
WEI S. Expression and clinical significance of serum MMP-9, MMP-2, TIMP-1, and TIMP-2 in infantile pneumonia[D]. Zunyi: Zunyi Medical University, 2023. (in Chinese)
[27] LI C, DENG T, CAO J, et al. Identifying ITGB2 as a potential prognostic biomarker in ovarian cancer[J]. Diagnostics (Basel, Switzerland), 2023, 13(6):1169.
[28] 贾红彦, 董静, 张宗德, 等. 结核分枝杆菌感染的免疫学检测技术研究进展及临床应用现状[J]. 中国防痨杂志，2022, 44(07): 720-6.
JIA H Y, DONG J, ZHANG Z D, et al. Advances in immunological detection techniques and clinical applications of Mycobacterium tuberculosis infection[J]. Chinese Journal of Antituberculosis, 2022, 44(7): 720-726. (in Chinese)
[29] 潘琳, 蔡睿志, 陶金, 等. IL-6经NF-κB信号通路上调人胎盘MSC SPP1表达促进M2型巨噬细胞极化的作用机制研究[J]. 细胞与分子免疫学杂志，2024, 40(11): 961-967.
PAN L, CAI R Z, TAO J, et al. IL-6 promotes M2 macrophage polarization via NF-κB signaling pathway by upregulating SPP1 expression in human placental MSCs[J]. Cellular & Molecular Immunology, 2024, 40(11): 961-967. (in Chinese)
[30] WANG C, LI Y, WANG L, et al. SPP1 represents a therapeutic target that promotes the progression of oesophageal squamous cell carcinoma by driving M2 macrophage infiltration[J]. Br J Cancer, 2024, 130(11): 1770-1782.
[31] KOGUCHI Y, KAWAKAMI K, UEZU K, et al. High plasma osteopontin level and its relationship with interleukin-12-mediated type 1 T helper cell response in tuberculosis[J]. Am J Respir Crit Care Med, 2003, 167(10): 1355-1359.
[32] NAU G J, LIAW L, CHUPP G L, et al. Attenuated host resistance against Mycobacterium bovis BCG infection in mice lacking osteopontin[J]. Infect Immun, 1999, 67(8): 4223-4230.
[33] 申福国, 杨钰, 孔维丽, 等. 肺结核病合并骨结核患者血清中KL-6和OPN表达及临床意义[J]. 中国防痨杂志，2024, 46(S1): 91-93.
SHEN F G, YANG Y, KONG W L, et al. Expression and clinical significance of serum KL-6 and OPN in pulmonary tuberculosis patients with bone tuberculosis[J]. Chinese Journal of Antituberculosis, 2024, 46(S1): 91-93. (in Chinese)
[34] 吴轶. 胸腔积液癌胚抗原与血清癌胚抗原比值和胸腔积液分泌性磷蛋白1在鉴别结核性胸腔积液和肺癌所致胸腔积液中的应用价值[J]. 广西医学，2020, 42(12): 1503-1506.
WU Y. Value of pleural fluid CEA/serum CEA ratio and pleural fluid SPP1 in differentiating tuberculous pleural effusion from lung cancer-related effusion[J]. Guangxi Medical Journal, 2020, 42(12): 1503-1506. (in Chinese)
[35] HATTORI T, IWASAKI-HOZUMI H, BAI G, et al. Both full-length and protease-cleaved products of osteopontin are elevated in infectious diseases[J]. Biomedicines, 2021, 9(8):1006.
[36] MAHMUD F J, DU Y, GREIF E, et al. Osteopontin/secreted phosphoprotein-1 behaves as a molecular brake regulating the neuroinflammatory response to chronic viral infection[J]. J Neuroinflammation, 2020, 17(1): 273.
[37] ARGANDONA LOPEZ C, BROWN A M. Microglial- neuronal crosstalk in chronic viral infection through mTOR, SPP1/OPN and inflammasome pathway signaling[J]. Front Immunol, 2024, 15: 1368465.
[38] 孙冰生，张真发. 骨桥蛋白及其受体CD44v对肿瘤侵袭调控的研究进展[J].中国肺癌杂志，2015，18 (11): 714-717.
SUN B S, ZHANG Z F. Advances in research of osteopontion and its receptor CD44v in tumor invasion and metastasis[J]. Chinese Journal of Lung Cancer, 2015, 18(11): 714-717. (in Chinese)
[39] 梁文婷, 霍开明, 古裕鸟, 等. 肺炎支原体肺炎患儿血清MICA、OPN水平及其与反复呼吸道感染的相关性研究[J]. 检验医学与临床，2024, 21(13): 1870-1874.
LIANG W T, HUO K M, GU Y N, et al. Correlation between serum MICA, OPN levels and recurrent respiratory infections in children with Mycoplasma pneumoniae pneumonia[J]. Laboratory Medicine and Clinical, 2024, 21(13): 1870-1874. (in Chinese)
[40] ICER M A, GEZMEN-KARADAG M. The multiple functions and mechanisms of osteopontin[J]. Clin Biochem, 2018, 59: 17-24.

Rv3435c重组耻垢分枝杆菌感染小鼠RAW264.7巨噬细胞的转录组分析

Transcriptome Analysis of RAW264.7 Macrophages Infected with Rv3435c Recombinant Mycobacterium smegmatis

PDF

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

编辑推荐

Metrics

本文评价

[1]	李佳鹏, 刘庆, 孙佳钰, 马泽芳, 崔凯. 基于转录组和蛋白质组分析筛选银黑狐毛色形成的关键基因[J]. 畜牧兽医学报, 2025, 56(9): 4379-4392.
[2]	刘添, 吴雨伦, 姚火春, 潘子豪. 副结核分枝杆菌三重TaqMan qPCR 检测方法的建立[J]. 畜牧兽医学报, 2025, 56(9): 4750-4758.
[3]	范婧, 李伟, 朱妍, 勿都巴拉, 史佳慧, 胡斯乐, 吴江鸿. 湖羊不同发育期瘤胃形态学变化及基因表达差异研究[J]. 畜牧兽医学报, 2025, 56(8): 3773-3786.
[4]	时文健, 徐磊, 张泽, 杨蕊, 辛凌翔, 王楠, 陈祥, 鑫婷. 基于结核分枝杆菌牛变种C68001随机突变体库筛选和鉴定成膜相关基因[J]. 畜牧兽医学报, 2025, 56(8): 3992-4006.
[5]	刘莎, 苏蒙, 高倩梅, 宋丹丽, 赵桂苹, 李建慧, 李庆贺. SIRT1基因激活后鸡巨噬细胞转录组分析[J]. 畜牧兽医学报, 2025, 56(6): 2661-2671.
[6]	刘子龙, 李乔, 吴怡, 王慧慧, 李讨讨, 马友记. 肝转录组揭示中草药饲料添加剂可能影响湖羊肝组织胆汁酸代谢和免疫功能[J]. 畜牧兽医学报, 2025, 56(6): 3014-3026.
[7]	朱海燕, 张菁怡, 晏雪勇, 梁海平, 魏庆, 曹际, 黄建珍. 基于转录组探究光周期对泰和乌鸡产蛋性能影响的分子机制[J]. 畜牧兽医学报, 2025, 56(5): 2123-2135.
[8]	余昕雅, 何海健, 王磊, 倪语晨, 杜静, 周莹珊, 董婉玉, 王晓杜. LncRNA 18850对猪流行性腹泻病毒复制的影响[J]. 畜牧兽医学报, 2025, 56(3): 1366-1375.
[9]	李常营, 李俊, 李锡锋, 毕师诚, 曹立亭. 基于转录组学探究日粮添加酵母β-葡聚糖对新城疫疫苗免疫鸡肠道免疫的影响[J]. 畜牧兽医学报, 2025, 56(3): 1441-1452.
[10]	苏蒙, 刘莎, 宋丹丽, 高倩梅, 郑麦青, 文杰, 赵桂苹, 李庆贺. 基于转录组测序筛选肉鸡腹水综合征相关候选基因[J]. 畜牧兽医学报, 2025, 56(2): 559-570.
[11]	胡瀚文, 图格琴, 任秀娟, 丁文淇, 宫文典, 贾紫洁, 史琳, 马木仁, 宝日格乐, 芒来, 白东义. 蒙古马两个类群肌纤维发育表型及基因表达谱比较研究[J]. 畜牧兽医学报, 2025, 56(2): 643-656.
[12]	吴双, 尹娜, 余莫涵, 平玉宇, 白皓, 陈世豪, 常国斌. TRIM39.2过表达对鸡巨噬细胞转录表达的影响[J]. 畜牧兽医学报, 2025, 56(1): 178-188.
[13]	鲁秀, 张名爱, 孔敏, 张晶, 王秉翰, 侯中一, 滕兴怡, 姜雅静, 凡文磊, 王宝维. 基于转录组和蛋白质组分析筛选五龙鹅产蛋相关候选基因[J]. 畜牧兽医学报, 2025, 56(1): 232-245.
[14]	王盛琪, 季新雨, 黄福青, 胡曼丽, 王柔淇, 耿玉欣, 孙迎雪, 齐智利, 张鑫. 添加红景天苷的全价粮对犬血液生化指标和肝转录组学的影响[J]. 畜牧兽医学报, 2025, 56(1): 455-465.
[15]	张肖旭, 李昊, 冯平捷, 杨豪, 李新月, 吕冉, 潘章源, 储明星. 单细胞转录组测序技术在家养动物中的应用[J]. 畜牧兽医学报, 2024, 55(8): 3276-3287.