猪多潜能干细胞系的研究进展

doi:10.16431/j.cnki.1671-7236.2023.01.007

Abstract

Abstract: Pig pluripotent stem cells are a kind of stem cells established in vitro with the capability of self-renewal and three germ layers differentiation, which can be used in developmental biology research, genome editing and disease modeling establishment.It has great value of application in animal husbandry production and regenerative medicine research.The culture system is of great significance for the successful construction of pluripotent stem cells, which contains a variety of components, including basic culture medium, amino acids and other nutrients, small molecular compounds (cytokines and signal pathway activators or inhibitors) to maintain cell pluripotency and inhibit its differentiation.There are many previous reports about porcine pluripotent stem cells, but effective culture system has not been obtained to maintain the long-term passage of porcine pluripotent stem cells and complete germline chimerism.Porcine pluripotent stem cells can be divided into three pluripotent states:Naïve state, Formative state and Primed state.And they can be classified into porcine expanded pluripotent stem cells, porcine embryonic stem cells, porcine pregastrulation stem cells and porcine induced pluripotent stem cells according to the derivation of the cell lines.Overall, the authors summarized the functional influences of current application of cytokines and signal pathway activators or inhibitors on the pluripotency and differentiation ability of porcine pluripotent stem cells.The present review could provide guidlines for the establishment of Naïve pluripotency porcine pluripotent stem cells with germline chimerism.

Key words: porcine; pluripotent stem cell lines; pluripotency

CLC Number:

S828

WANG Zhilong, LIU Li, ZHANG Lu, GAO Shuai. Research Progress of Porcine Pluripotent Stem Cell Lines[J]. China Animal Husbandry & Veterinary Medicine, 2023, 50(1): 67-75.

References

[1] KUMAR D, TALLURI T R, SELOKAR N L, et al.Perspectives of pluripotent stem cells in livestock[J].World Journal Stem Cells, 2021, 13(1):1-29.
[2] ALBERIO R, CROXALL N, ALLEGRUCCI C.Pig epiblast stem cells depend on activin/nodal signaling for pluripotency and self-renewal[J]. Stem Cells Development, 2010, 19(10):1627-1636.
[3] KOBAYASHI T, ZHANG H, TANG W W C, et al.Principles of early human development and germ cell program from conserved model systems[J].Nature, 2017, 546(7658):416-420.
[4] NIU D, WEI H J, LIN L, et al.Inactivation of porcine endogenous Retrovirus in pigs using CRISPR-Cas9[J]. Science, 2017, 357(6357):1303-1307.
[5] YUE Y, XU W, KAN Y, et al.Extensive germline genome engineering in pigs[J].Nature Biomedical Engineering, 2021, 5(2):134-143.
[6] YAN S, TU Z, LIU Z, et al.A Huntingtin knockin pig model recapitulates features of selective neurodegeneration in Huntington's disease[J]. Cell, 2018, 173(4):989-1002.e1013.
[7] 崔文涛, 单同领, 李奎.转基因猪的研究现状及应用前景[J] 中国畜牧兽医, 2007, 34(4):58-62. CUI W T, SHAN T L, LI K.Research status and application prospects of transgenic pigs[J].China Animal Husbandry & Veterinary Medicine, 2007, 34(4):58-62.(in Chinese)
[8] HACKETT JA, SURANI MA.Regulatory principles of pluripotency:From the ground state up[J].Cell Stem Cell, 2014, 15(4):416-430.
[9] NICHOLS J, SMITH A.Naive and primed pluripotent states[J].Cell Stem Cell, 2009, 4(6):487-492.
[10] Smith A.Formative pluripotency:The executive phase in a developmental continuum[J].Development, 2017, 144(3):365-373.
[11] YU L, WEI Y, SUN HX, et al.Derivation of intermediate pluripotent stem cells amenable to primordial germ cell specification[J].Cell Stem Cell, 2021, 28(3):550-567.
[12] YING Q L, WRAY J, NICHOLS J, et al.The ground state of embryonic stem cell self-renewal[J].Nature, 2008, 453(7194):519-523.
[13] ROSSANT J.Stem cells and early lineage development[J].Cell, 2008, 132(4):527-531.
[14] BRADLEY A, EVANS M, KAUFMAN M H, et al.Formation of germ-line chimaeras from embryo-derived teratocarcinoma cell lines[J]. Nature, 1984, 309(5965):255-256.
[15] NAGY A, GÓCZA E, DIAZ E M, et al.Embryonic stem cells alone are able to support fetal development in the mouse[J].Development, 1990, 110(3):815-821.
[16] TESAR P J, CHENOWETH J G, BROOK F A, et al.New cell lines from mouse epiblast share defining features with human embryonic stem cells[J].Nature, 2007, 448(7150):196-199.
[17] BRONS I G, SMITHERS L E, TROTTER M W, et al.Derivation of pluripotent epiblast stem cells from mammalian embryos[J].Nature, 2007, 448(7150):191-195.
[18] TACHIBANA M, SPARMAN M, RAMSEY C, et al.Generation of chimeric rhesus monkeys[J].Cell, 2012, 148(1-2):285-295.
[19] HUANG Y, OSORNO R, TSAKIRIDIS A, et al.In vivo differentiation potential of epiblast stem cells revealed by chimeric embryo formation[J]. Cell Reports, 2012, 2(6):1571-1578.
[20] KALKAN T, SMITH A.Mapping the route from naive pluripotency to lineage specification[J].Philosophical Transactions of the Royal Society B:Biological Sciences, 2014, 369(1657):20130540.
[21] WANG X, XIANG Y, YU Y, et al.Formative pluripotent stem cells show features of epiblast cells poised for gastrulation[J]. Cell Research, 2021, 31(5):526-541.
[22] KINOSHITA M, BARBER M, MANSFIELD W, et al.Capture of mouse and human stem cells with features of formative pluripotency[J].Cell Stem Cell, 2021, 28(12):2180.
[23] MORGANI S, NICHOLS J, HADJANTONAKIS A K.The many faces of pluripotency:In vitro adaptations of a continuum of in vivo states[J].BMC Developmental Biology, 2017, 17(1):7.
[24] TANG F, BARBACIORU C, BAO S, et al.Tracing the derivation of embryonic stem cells from the inner cell mass by single-cell RNA-Seq analysis[J].Cell Stem Cell, 2010, 6(5):468-478.
[25] YAN L, YANG M, GUO H, et al.Single-cell RNA-Seq profiling of human preimplantation embryos and embryonic stem cells[J]. Nature Structural & Molecular Biology, 2013, 20(9):1131-1139.
[26] NAKAMURA T, OKAMOTO I, SASAKI K, et al.A developmental coordinate of pluripotency among mice, monkeys and humans[J]. Nature, 2016, 537(7618):57-62.
[27] PETROPOULOS S, EDSGARD D, REINIUS B, et al.Single-cell RNA-Seq reveals lineage and X chromosome dynamics in human preimplantation embryos[J].Cell, 2016, 167(1):285.
[28] PIJUAN-SALA B, GRIFFITHS J A, GUIBENTIF C, et al.A single-cell molecular map of mouse gastrulation and early organogenesis[J]. Nature, 2019, 566(7745):490-495.
[29] RAMOS-IBEAS P, SANG F, ZHU Q, et al.Pluripotency and X chromosome dynamics revealed in pig pre-gastrulating embryos by single cell analysis[J]. Nature Communications, 2019, 10(1):500.
[30] LIU T, LI J, YU L, et al.Cross-species single-cell transcriptomic analysis reveals pre-gastrulation developmental differences among pigs, monkeys, and humans[J]. Cell Discovery, 2021, 7(1):14.
[31] CAO S, HAN J, WU J, et al.Specific gene-regulation networks during the pre-implantation development of the pig embryo as revealed by deep sequencing[J]. BMC Genomics, 2014, 15(1):4.
[32] WEI Q, LI R, ZHONG L, et al.Lineage specification revealed by single-cell gene expression analysis in porcine preimplantation embryos[J]. Biology of Reproduction, 2018, 99(2):283-292.
[33] ZHI M L, ZHANG J Y, TANG Q Z, et al.Generation and characterization of stable pig pregastrulation epiblast stem cell lines[J]. Cell Research, 2021, 32(4):383-400.
[34] ZHOU Y, ZHOU B, PACHE L, et al.Metascape provides a biologist-oriented resource for the analysis of systems-level datasets[J].Nature Communications, 2019, 10(1):1523.
[35] BLAKELEY P, FOGARTY N M, DEL VALLE I, et al.Defining the three cell lineages of the human blastocyst by single-cell RNA-Seq[J]. Development, 2015, 142(18):3151-3165.
[36] PIEDRAHITA J A, ANDERSON G B, BONDURANT R H.Influence of feeder layer type on the efficiency of isolation of porcine embryo-derived cell lines[J].Theriogenology, 1990, 34(5):865-877.
[37] STROJEK R M, REED M A, HOOVER J L, et al.A method for cultivating morphologically undifferentiated embryonic stem cells from porcine blastocysts[J].Theriogenology, 1990, 33(4):901-913.
[38] YANG J, RYAN D J, WANG W, et al.Establishment of mouse expanded potential stem cells[J]. Nature, 2017, 550(7676):393-397.
[39] YANG J, RYAN D J, LAN G, et al.In vitro establishment of expanded-potential stem cells from mouse pre-implantation embryos or embryonic stem cells[J].Nature Protocols, 2019, 14(2):350-378.
[40] GAO X, NOWAK-IMIALEK M, CHEN X, et al.Establishment of porcine and human expanded potential stem cells[J]. Nature Cell Biology, 2019, 21(6):687-699.
[41] ZHAO L, GAO X, ZHENG Y, et al.Establishment of bovine expanded potential stem cells[J].Proceedings of the National Academy of Sciences of the United States of America, 2021, 118(15):e2018505118.
[42] AZZOLIN L, PANCIERA T, SOLIGO S, et al.YAP/TAZ incorporation in the β-catenin destruction complex orchestrates the Wnt response[J]. Cell, 2014, 158(1):157-170.
[43] WANG W, LI N, LI X, et al.Tankyrase inhibitors target YAP by stabilizing angiomotin family proteins[J].Cell Reports, 2015, 13(3):524-532.
[44] MARTIN G R.Isolation of a pluripotent cell line from early mouse embryos cultured in medium conditioned by teratocarcinoma stem cells[J].Proceedings of the National Academy of Sciences of the United States of America, 1981, 78(12):7634-7638.
[45] BUEHR M, MEEK S, BLAIR K, et al.Capture of authentic embryonic stem cells from rat blastocysts[J].Cell, 2008, 135(7):1287-1298.
[46] THOMSON J A, ITSKOVITZ-ELDOR J, SHAPIRO S S, et al.Embryonic stem cell lines derived from human blastocysts[J]. Science, 1998, 282(5391):1145-1147.
[47] THOMSON J A, KALISHMAN J, GOLOS T G, et al.Isolation of a primate embryonic stem cell line[J].Proceedings of the National Academy of Sciences of the United States of America, 1995, 92(17):7844-7848.
[48] NAVARRO M, SOTO D A, PINZON C A, et al.Livestock pluripotency is finally captured in vitro[J].Reproduction, Fertility and Development, 2019, 32(2):11-39.
[49] PANASOPHONKUL S, THARASANIT T, TECHAKUMPHU M.Establishment of porcine embryonic stem-like cells from parthenogenetic and in vivo derived embryos[J]. The Thai Journal of Veterinary Medicine, 2010, 40(3):273-280
[50] 邓守龙, 彭涛, 吕自力, 等.哺乳动物胚胎干细胞研究进展[J].中国畜牧兽医, 2009, 36(9):86-93. DENG S L, PENG T, LYU Z L, et al.Mammalian embryo stem cell research progress[J].China Animal Husbandry & Veterinary Medicine, 2009, 36(9):86-93.(in Chinese)
[51] 王朋朋, 黄书林, 张云科, 等.哺乳动物细胞无血清全悬浮培养技术研究进展[J].中国畜牧兽医, 2021, 48(3):839-845. WANG P P, HUANG S L, ZHANG Y K, et al.Research progress on serum-free suspension culture of mammalian cells[J].China Animal Husbandry & Veterinary Medicine, 2021, 48(3):839-845.(in Chinese)
[52] WANG J, GU Q, HAO J, et al.Tbx3 and Nr5α2 play important roles in pig pluripotent stem cells[J].Stem Cell Reviews and Reports, 2013, 9(5):700-708.
[53] XUE B, LI Y, HE Y, et al.Porcine pluripotent stem cells derived from IVF embryos contribute to chimeric development in vivo[J].PLoS One, 2016, 11(3):e0151737.
[54] VASSILIEV I, VASSILIEVA S, BEEBE L F, et al.In vitro and in vivo characterization of putative porcine embryonic stem cells[J].Cell Reprogram, 2010, 12(2):223-230.
[55] GAO X F, NOWAK-IMIALEK M, CHEN X, et al.Establishment of porcine and human expanded potential stem cells[J]. Nature Cell Biology, 2019, 21(6):687-699.
[56] YUAN Y, PARK J, TIAN Y, et al.A six-inhibitor culture medium for improving naïve-type pluripotency of porcine pluripotent stem cells[J]. Cell Death Discovery, 2019, 5:104.
[57] CHOI K H, LEE D K, KIM S W, et al.Chemically defined media can maintain pig pluripotency network in vitro[J]. Stem Cell Reports, 2019, 13(1):221-234.
[58] HOU D R, JIN Y, NIE X W, et al.Derivation of porcine embryonic stem-like cells from in vitro-produced blastocyst-stage embryos[J].Scientific Reports, 2016, 6:25838.
[59] BREVINI T A, PENNAROSSA G, ATTANASIO L, et al.Culture conditions and signalling networks promoting the establishment of cell lines from parthenogenetic and biparental pig embryos[J].Stem Cell Reviews and Reports, 2010, 6(3):484-495.
[60] HARAGUCHI S, KIKUCHI K, NAKAI M, et al.Establishment of self-renewing porcine embryonic stem cell-like cells by signal inhibition[J].Journal of Reproduction and Development, 2012, 58(6):707-716.
[61] PARK J K, KIM H S, UH K J, et al.Primed pluripotent cell lines derived from various embryonic origins and somatic cells in pig[J]. PLoS One, 2013, 8(1):e52481.
[62] MA Y, YU T, CAI Y, et al.Preserving self-renewal of porcine pluripotent stem cells in serum-free 3i culture condition and independent of LIF and b-FGF cytokines[J]. Cell Death Discovery, 2018, 4:21.
[63] KIM H, WU J, YE S, et al.Modulation of beta-catenin function maintains mouse epiblast stem cell and human embryonic stem cell self-renewal[J].Nature Communications, 2013, 4:2403.
[64] GREBER B, WU G, BERNEMANN C, et al.Conserved and divergent roles of FGF signaling in mouse epiblast stem cells and human embryonic stem cells[J]. Cell Stem Cell, 2010, 6(3):215-226.
[65] VAN OOSTEN A L, COSTA Y, SMITH A, et al.JAK/STAT3 signalling is sufficient and dominant over antagonistic cues for the establishment of naive pluripotency[J]. Nature Communications, 2012, 3:817.
[66] LI X, ZHU L, YANG A, et al.Calcineurin-NFAT signaling critically regulates early lineage specification in mouse embryonic stem cells and embryos[J].Cell Stem Cell, 2011, 8(1):46-58.
[67] YANG Y, LIU B, XU J, et al.Derivation of pluripotent stem cells with in vivo embryonic and extraembryonic potency[J].Cell, 2017, 169(2):243-257.e225.
[68] TAKAHASHI K, YAMANAKA S.Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors[J].Cell, 2006, 126(4):663-676.
[69] YU J, VODYANIK M A, SMUGA-OTTO K, et al.Induced pluripotent stem cell lines derived from human somatic cells[J].Science, 2007, 318(5858):1917-1920.
[70] TAKAHASHI K, TANABE K, OHNUKI M, et al.Induction of pluripotent stem cells from adult human fibroblasts by defined factors[J].Cell, 2007, 131(5):861-872.
[71] WEST F D, TERLOUW S L, KWON D J, et al.Porcine induced pluripotent stem cells produce chimeric offspring[J]. Stem Cells and Development, 2010, 19(8):1211-1220.
[72] CHENG D, GUO Y, LI Z, et al.Porcine induced pluripotent stem cells require LIF and maintain their developmental potential in early stage of embryos[J].PLoS One, 2012, 7(12):e51778.
[73] FUJISHIRO S H, NAKANO K, MIZUKAMI Y, et al.Generation of naive-like porcine-induced pluripotent stem cells capable of contributing to embryonic and fetal development[J].Stem Cells and Development, 2013, 22(3):473-482.
[74] PETKOV S, GLAGE S, NOWAK-IMIALEK M, et al.Long-term culture of porcine induced pluripotent stem-like cells under feeder-free conditions in the presence of histone deacetylase inhibitors[J].Stem Cells and Development, 2016, 25(5):386-394.
[75] SECHER J O, CEYLAN A, MAZZONI G, et al.Systematic in vitro and in vivo characterization of leukemia-inhibiting factor- and fibroblast growth factor-derived porcine induced pluripotent stem cells[J]. Molecular Reproduction and Development, 2017, 84(3):229-245.
[76] ZHANG W, WANG H, ZHANG S, et al.Lipid supplement in the cultural condition facilitates the porcine iPSC derivation through cAMP/PKA/CREB signal pathway[J].International Journal of Molecular Sciences, 2018, 19(2):509.
[77] LI D, SECHER J, HYTTEL P, et al.Generation of transgene-free porcine intermediate type induced pluripotent stem cells[J].Cell Cycle, 2018, 17(23):2547-2563.
[78] THOMSON A J, PIERART H, MEEK S, et al.Reprogramming pig fetal fibroblasts reveals a functional LIF signaling pathway[J].Cell Reprogram, 2012, 14(2):112-122.
[79] LIU K, JI G, MAO J, et al.Generation of porcine-induced pluripotent stem cells by using Oct4 and Klf4 porcine factors[J].Cell Reprogram, 2012, 14(6):505-513.
[80] CHAKRITBUDSABONG W, SARIYA L, PAMONSUPORNVICHIT S, et al.Generation of a pig induced pluripotent stem cell (piPSC) line from embryonic fibroblasts by incorporating LIN28 to the four transcriptional factor-mediated reprogramming:VSMUi001-D[J].Stem Cell Research, 2017, 24:21-24.
[81] KUES W A, HERRMANN D, BARG-KUES B, et al.Derivation and characterization of sleeping beauty transposon-mediated porcine induced pluripotent stem cells[J].Stem Cells and Development, 2013, 22(1):124-135.
[82] 廖荣荣, 颉孝贤, 马育芳, 等.诱导多能干细胞技术在猪转基因研究中的应用前景[J]. 中国畜牧兽医, 2013, 40(6):90-95. LIAO R R, XIE X X, MA Y F, et al.The application prospect of induced pluripotent stem cells in pig transgenic research[J].China Animal Husbandry & Veterinary Medicine, 2013, 40(6):90-95.(in Chinese)

Research Progress of Porcine Pluripotent Stem Cell Lines

PDF (PC)

Knowledge

Abstract

Cite this article

share this article

References

Related Articles 15

Recommended Articles

Metrics

Comments

[1]	CHEN Zhian, ZHANG Beiwen, HE Minjia, CHEN Meichun, WENG Chengzhen, HUANG Xinxin, LI Hongxi, ZENG Zhongwen, CHEN Baoliang, QIU Longxin, CHEN Hongbo, LI Xiaobing. Genetic Variation and Codon Usage Bias Analysis of Porcine Pseudorabies Virus gE Gene [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(7): 3264-3275.
[2]	ZHANG Limeng, LI Runting, SONG Yue, NIE Xiaoning, KONG Li, SHAN Jingwei, XU Yingying, WANG Linqing, CHEN Longxin. Construction and Screening of Single Chain Antibody Library of Porcine Parvovirus NS1 Protein [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(7): 3276-3285.
[3]	RUAN Shihui, LIU Chunyan, WEI Yangyang, HE Yiyi, WU Qiwen, XIONG Yunxia, YANG Xuefen, WANG Li, YI Hongbo. Effect of PDCoV on the Extracellular Matrix of the Intestine and Its Dynamic Changes in Weaned Piglets [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(7): 3297-3307.
[4]	ZHANG Bingyan, FAN Rui, FENG Shutang, JIA Junting, ZHANG Jianbin, MA Yuyuan. Whole Genome Resequencing Analysis of PERV Non-transmitting Zhong Xu Wuzhishan Mini-pig Inbred Line [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(6): 2459-2467.
[5]	WEI Huangsiwu, ZHANG Xingyi, HUANG Xiaohua, LIU Changjin, WU Wenjie, SHEN Zhengqiao, LUO Feng, DENG Shunzhou. Eukaryotic Expression of Porcine Rotavirus VP6 Protein and Preparation and Application of Its Monoclonal Antibody [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(6): 2750-2761.
[6]	ZHONG Weilin, YAO Jianhui, WEI Xiaoqi, YAN Guangzhi, CHEN Shengnan, LIU Mingjie, HUANG Liangzong. Diagnosis and Genetic Evolution Analysis of PCV2d Infection in a Pig Farm in Guangdong [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(6): 2781-2789.
[7]	HUANG Xiaojiu, LEI Lei, PENG Xiaoye, WANG Kaixin, CHEN Yingyi, WANG Jixian, WANG Yuge, DUAN Deyong, YANG Yi, WANG Aibing. Construction of a IPEC-J2 Cell Line Stably Overexpressing NM-ⅡA Tail and Its Effect on Porcine Epidemic Diarrhea Virus Infection [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(5): 2243-2252.
[8]	JIAO Xianqin, MA Xiao, TIAN Runbo, LIU Ying, MA Shijie, CHEN Hongying. Construction of Recombinant Porcine Pseudorabies Virus Expressing Cap Protein of Porcine Circovirus Type 2d [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(5): 2278-2286.
[9]	PAN Xiangying, ZENG Zhiyong, LIANG Haiying, TANG Deyuan, WANG Bin, YE Ni, TIAN Hongli, BIAN Mengting, LIU Jiajia, HUANG Shu. Truncated Expression of Porcine Rotavirus VP6 Protein and Establishment of an Indirect ELISA Antibody Detection Method [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(3): 1231-1240.
[10]	MAO Fuchao, ZHAI Chongkai, TIAN Wenjing, WANG Conghui, SONG Minjie, WANG Yingxian, ZHANG Hewei. An Updated Review of Porcine Deltacoronavirus in Terms of Infection and Anti-infection Research [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(3): 1281-1291.
[11]	LI Huanhuan, YU Chenmin, TIAN Xiaorong, LI Rui, LI Zongyun, ZHANG Yanyan, ZHAO Di, WANG Lei, HOU Yongqing, WU Tao. Protective Effect of Catechu Extract on the Colon of Young Piglets Infected with Porcine Epidemic Diarrhea Virus [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(3): 1360-1369.
[12]	YANG Li, DU Xiaomei, LIU Mengyuan, WU Shenglong, BAO Wenbin, WU Zhengchang. Screening and Functional Verification of Key lncRNA Affecting Porcine Epidemic Diarrhea Virus Replication [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(2): 792-800.
[13]	BAI Yihan, WANG Dongliang. Research Progress on Genetic and Host Diversity of Porcine Circoviruses [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(2): 832-843.
[14]	XIANG Jiaojiao, YUAN Na, LI Huihui, SHAO Mingzhu, ZHAO Fuping, ZHANG Longchao, WANG Lixian, SHI Lijun, CHEN Bin. Function Study on SOX12 Gene in Vero Cells Infected with Porcine Epidemic Diarrhea Virus [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(1): 289-297.
[15]	LI Songbei, XIE Yongsheng, LONG Xiaoqin, ZHANG Xiaoxiao, CHEN Yongjie, ZHANG Chunhong, JIAO Maoxing, GUO Chunhe. Genetic Variation Analysis of Porcine Reproductive and Respiratory Syndrome Virus in South China in 2023 [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(1): 331-340.