Abstract:Gene editing technology provides a new solution for pig (Sus scrofa) molecular breeding. Gene editing technology can achieve accurate and efficient gene editing, providing a powerful tool for breeding pig. Gene editing can not only be used to quickly and accurately improve growth rate and disease resistance, thereby enhancing the feeding efficiency of pigs, but also optimize the quality, taste and even nutritional content of pork to meet the growing consumer demand for high-quality, healthy pork. However, there are still many technical challenges in the application of gene editing technology, such as off-target effects. It is necessary to increase technological innovation to solve these problems restricting the efficiency of pig gene editing breeding, and promote the wide application of gene editing technology in pig disease resistance breeding. This paper summarized the latest development of gene editing technology in pig molecular breeding, including technical progress, application status and challenges. This review provides reference for breeding of disease resistance in China.
[1] Andersen O M, Bogh N, Landau A M, et al.2022. A genetically modified minipig model for Alzheimer's disease with sorl1 haploinsufficiency[J]. Cell Reports Medicine, 3(9): 100740. [2] Bazaz M, Adeli A, Azizi M, et al.2022. Recent developments in miRNA based recombinant protein expression in CHO[J]. Biotechnology Letters, 44(5-6): 671-681. [3] Beslika E, Leite-Moreira A, De Windt L J, et al.2024. Large animal models of pressure overload-induced cardiac left ventricular hypertrophy to study remodelling of the human heart with aortic stenosis[J]. Cardiovascular Research, 120(5): 461-475. [4] Biagioni A, Laurenzana A, Margheri F, et al.2019. Erratum: Publisher correction to: Delivery systems of CRISPR/Cas9-based cancer gene therapy[J]. Journal of Biological Engineering, 13: 37. [5] Burkard C, Opriessnig T, Mileham A J, et al.2018. Pigs lacking the scavenger receptor cysteine-rich domain 5 of Cd163 are resistant to Porcine reproductive and respiratory syndrome virus 1 infection[J]. Journal of Virology, 92(16). [6] Carey K, Ryu J, Uh K, et al.2019. Frequency of off-targeting in genome edited pigs produced via direct injection of the CRISPR/Cas9 system into developing embryos[J]. BMC Biotechnology, 19(1): 25. [7] Chen P R, Rowland R R R, Stoian A M, et al.2022. Disruption of anthrax toxin receptor 1 in pigs leads to a rare disease phenotype and protection from Senecavirus A infection[J]. Scientific Reports, 12(1): 5009. [8] Chen Z H, Yuan F F, Li Y H, et al.2016. Construction and characterization of a full-length cdna infectious clone of emerging porcine Senecavirus A[J]. Virology, 497: 111-124. [9] Cromwell C R, Sung K, Park J, et al.2018. Incorporation of bridged nucleic acids into CRISPR rnas improves Cas9 endonuclease specificity[J]. Nature Communications, 9(1): 1448. [10] Cuesta-Geijo M A, Garcia-Dorival I, Del Puerto A, et al.2022. New insights into the role of endosomal proteins for African swine fever virus infection[J]. PLOS Pathogens, 18(1): e1009784. [11] Delmas B B, Gelfi J, L'Haridon R, et al.1992. Aminopeptidase N is a major receptor for the entero-pathogenic coronavirus TGEV[J]. Nature, 357(6377):417-420. [12] Gao F, Li P, Yin Y, et al.2023. Molecular breeding of livestock for disease resistance[J]. Virology, 587: 109862. [13] Gao Q, Yang Y, Luo Y, et al.2022. Adaptation of African swine fever virus to porcine kidney cells stably expressing Cd163 and SIGLEC1[J]. Frontires in Immunology, 13: 1015224. [14] Geisert R D, Johns D N, Pfeiffer C A, et al.2023. Gene editing provides a tool to investigate genes involved in reproduction of pigs[J]. Molecular Reproduction and Development, 90(7): 459-468. [15] Golovan S P, Meidinger R G, Ajakaiye A, et al.2001. Pigs expressing salivary phytase produce low-phosphorus manure[J]. Nature Biotechnology, 19(8): 741-745. [16] Guo C H, Wang M, Zhu Z B, et al.2019. Highly efficient generation of pigs harboring a partial deletion of the Cd163 SRCR5 domain, which are fully resistant to Porcine reproductive and respiratory syndrome virus 2 infection[J]. Fronters in Immunology, 10: 1846. [17] Ji C M, Wang B, Zhou J, et al.2018. Aminopeptidase-N-independent entry of Porcine epidemic diarrhea virus into VERO or porcine small intestine epithelial cells[J]. Virology, 517: 16-23. [18] Joshi L R, Mohr K A, Clement T, et al.2016. Detection of the emerging Picornavirus Senecavirus A in pigs, mice, and houseflies[J]. Journal of Clinical Microbiology, 54(6): 1536-1545. [19] Jung I Y, Lee J.2018. Unleashing the therapeutic potential of CAR-T cell therapy using gene-editing technologies[J]. Molecules and Cells, 41(8): 717-723. [20] Klinkovskij A, Shepelev M, Isaakyan Y, et al.2023. Advances of genome editing with CRISPR/Cas9 in neurodegeneration: The right path towards therapy[J]. Biomedicines, 11(12): 3333. [21] Lamas-Toranzo I, Guerrero-Sanchez J, Miralles-Bover H, et al.2017. CRISPR is knocking on barn door[J]. Reproduction in Domestic Animals, 52 Suppl 4: 39-47. [22] Li R Q, Zeng W, Ma M, et al.2020. Precise editing of myostatin signal peptide by CRISPR/Cas9 increases the muscle mass of Liang Guang Small Spotted pigs[J]. Transgenic Research, 29(1): 149-163. [23] Li W, Hulswit Rjg, Kenney S P, et al.2018. Broad receptor engagement of an emerging global Coronavirus may potentiate its diverse cross-species transmissibility[J]. Proceedings of the National Academy of Sciences of the USA, 115(22): E5135-E5143. [24] Li Y Y, Wang H, Chen H Y, et al.2022. Generation of a genetically modified pig model with CREBRFR457Q variant[J]. FASEB Journal, 36(11): e22611. [25] Lin J, Cao C W, Tao C, et al.2017. Cold adaptation in pigs depends on UCP3 in beige adipocytes[J]. Journal of Molecular Cell Biology, 9(5): 364-375. [26] Liu X F, Liu H B, Wang M, et al.2019. Disruption of the ZBED6 binding site in intron 3 of IGF2 by CRISPR/Cas9 leads to enhanced muscle development in liang guang small spotted pigs[J]. Transgenic Research, 28(1): 141-150. [27] Liu Z Y, Zhang M J, Huang P X, et al.2023. Generation of apn-chimeric gene-edited pigs by CRISPR/Cas9-mediated knock-in strategy[J]. Gene, 851: 147007. [28] Llobat L.2020. Embryo gene expression in pig pregnancy[J]. Reproduction in Domestic Animals, 55(4): 523-529. [29] Mcgaugh S, Schwartz T S.2017. Here and there, but not everywhere: repeated loss of uncoupling protein 1 in amniotes[J]. Biology Letters (2005), 13(1): 20160749. [30] Meyer A E, Pfeiffer C A, Brooks K E, et al.2019. New perspective on conceptus estrogens in maternal recognition and pregnancy establishment in the pigdagger[J]. Biology of Reproduction, 101(1): 148-161. [31] Palgrave C J, Gilmour L, Lowden C S, et al.2011. Species-specific variation in RELA underlies differences in NF-κB activity: A potential role in African swine fever pathogenesis[J]. Journal of Virology, 85(12): 6008-6014. [32] Popescu L, Gaudreault N N, Whitworth K M, et al.2017. Genetically edited pigs lacking Cd163 show no resistance following infection with the African swine fever virus isolate, georgia 2007/1[J]. Virology, 501: 102-106. [33] Prather R S, Rowland R R, Ewen C, et al.2013. An intact sialoadhesin (SN/SIGLEC1/CD169) is not required for attachment/internalization of the Porcine reproductive and respiratory syndrome virus[J]. Journal of Virology, 87(17): 9538-9546. [34] Qi C Y, Pang D X, Yang K, et al.2022. Generation of PCBP1-deficient pigs using CRISPR/Cas9-mediated gene editing[J]. Iscience, 25(10): 105268. [35] Qi Y Y, Zhang Y, Tian S J, et al.2023. An optimized prime editing system for efficient modification of the pig genome[J]. Science China-Life Sciences, 66(12): 2851-2861. [36] Ren J L, Hai T, Chen Y C, et al.2024. Improve meat production and virus resistance by simultaneously editing multiple genes in livestock using Cas12iMax[J]. Science China-Life Sciences, 67(3): 555-564. [37] Ricroch A.2019. Global developments of genome editing in agriculture[J]. Transgenic Research, 28(Suppl 2): 45-52. [38] Sánchez-Torres T C, Gomez P P, Gomez D M, et al.2003. Expression of porcine Cd163 on monocytes/macrophages correlates with permissiveness to african swine fever infection[J]. Archives of Virology, 148(12): 2307-2323. [39] Stoian A M M, Rowland R R R, Brandariz-Nunez A.2022. Mutations within scavenger receptor cysteine-rich (SRCR) protein domain 5 of porcine CD163 involved in infection with Porcine reproductive and respiratory syndrome virus (PRRS)[J]. Journal of General Virology, 103(5): 1740. [40] Stoian A M M, Rowland R R R, Petrovan V, et al.2020. The use of cells from ANPEP knockout pigs to evaluate the role of aminopeptidase N (APN) as a receptor for Porcine deltacoronavirus (PDCoV)[J]. Virology, 541: 136-140. [41] Sun L M, Zhao C Z, Fu Z, et al.2021. Genome-scale CRISPR screen identifies TMEM41B as a multi-function host factor required for Coronavirus replication[J]. Plos Pathogens, 17(12): e1010113. [42] Tang Y D, Liu J T, Wang T Y, et al.2017. Crispr/cas9-mediated multiple single guide RNAS potently abrogate Pseudorabies virus replication[J]. Archives of Virology, 162(12): 3881-3886. [43] Tu C F, Chuang C K, Hsiao K H, et al.2019. Lessening of Porcine epidemic diarrhoea virus susceptibility in piglets after editing of the CMP-N-glycolylneuraminic acid hydroxylase gene with CRISPR/Cas9 to nullify N-glycolylneuraminic acid expression[J]. PLOS ONE, 14(5): e217236. [44] Tu C F, Chuang C K, Yang T S.2022. The application of new breeding technology based on gene editing in pig industry - A review[J]. Animal Bioscience, 35(6): 791-803. [45] Tu C F, Peng S H, Chuang C K, et al.2023. Reproductive technologies needed for the generation of precise gene-edited pigs in the pathways from laboratory to farm[J]. Animal Bioscience, 36(2): 339-349. [46] Van Breedam W, Delputte P L, Van G H, et al.2010. Porcine reproductive and respiratory syndrome virus entry into the porcine macrophage[J]. Journal of General Virology, 91(Pt 7): 1659-1667. [47] Wang B, Liu Y, Ji C M, et al.2018. Porcine deltacoronavirus engages the transmissible gastroenteritis virus functional receptor porcine aminopeptidase N for infectious cellular entry[J]. Journal of Virology, 92(12). [48] Wang K K, Ouyang H S, Xie Z C, et al.2015. Efficient generation of myostatin mutations in pigs using the CRISPR/Cas9 system[J]. Scientific Reports, 5: 16623. [49] Wang K K, Tang X C, Xie Z C, et al.2017. Crispr/Cas9-mediated knockout of myostatin in chinese indigenous erhualian pigs[J]. Transgenic Research, 26(6): 799-805. [50] Wells K D, Bardot R, Whitworth K M, et al.2017. Replacement of porcine Cd163 scavenger receptor cysteine-rich domain 5 with a Cd163-like homolog confers resistance of pigs to genotype 1 but not genotype 2 Porcine reproductive and respiratory syndrome virus[J]. Journal of Virology, 91(2). [51] Whyte J J, Meyer A E, Spate L D, et al.2018. Inactivation of porcine interleukin-1beta results in failure of rapid conceptus elongation[J]. Proceedings of the National Academy of Sciences of the USA, 115(2): 307-312. [52] Xiang G H, Ren J L, Hai T, et al.2018. Editing porcine IGF2 regulatory element improved meat production in chinese bama pigs[J]. Cellular and Molecular Life Sciences, 75(24): 4619-4628. [53] Xie Z C, Jiao H P, Xiao H N, et al.2020. Generation of PRSAD2 gene knock-in pig via CRISPR/Cas9 technology[J]. Antiviral Research, 174: 104696. [54] Xie Z C, Pang D X, Yuan H M, et al.2018. Genetically modified pigs are protected from classical Swine fever virus[J]. PLOS Pathogens, 14(12): e1007193. [55] Xu K, Zhou Y R, Mu Y L, et al.2020. Cd163 and pAPN double-knockout pigs are resistant to PRRSV and TGEV and exhibit decreased susceptibility to PDCoV while maintaining normal production performance[J]. Elife, 9:e57132. [56] Xu K, Zhou Y R, Shang H T, et al.2023. Pig macrophages with site-specific edited Cd163 decrease the susceptibility to infection with Porcine reproductive and respiratory syndrome virus[J]. Journal of Integrative Agriculture, 22(7): 2188-2199. [57] Yang H Q, Wu Z F.2018. Genome editing of pigs for agriculture and biomedicine[J]. Fronters in Genetics, 9: 360. [58] Yang H Q, Zhang J, Zhang X W, et al.2018. Cd163 knockout pigs are fully resistant to highly pathogenic Porcine reproductive and respiratory syndrome virus[J]. Antiviral Research, 151: 63-70. [59] Yao J, Zeng H S, Zhang M, et al.2019. OSBPL2-disrupted pigs recapitulate dual features of human hearing loss and hypercholesterolaemia[J]. Journal of Genetics and Genomics, 46(8): 379-387. [60] Yuan T L, Wu L L, Li S Y, et al.2024. Deep learning models incorporating endogenous factors beyond DNA sequences improve the prediction accuracy of base editing outcomes[J]. Cell Discovery, 10(1): 20. [61] Zhang F L, Fu Y M, Wang J F, et al.2024. Conjugated linoleic acid (CLA) reduces HFD-induced obesity by enhancing bat thermogenesis and iWAT browning via the CD36-AMPK pathway[J]. Cell Biochemistry and Function, 42(2): e3937. [62] Zhang J F, Khazalwa E M, Abkallo H M, et al.2021. The advancements, challenges, and future implications of the CRISPR/Cas9 system in swine research[J]. Journal of Genetics and Genomics, 48(5): 347-360. [63] Zhao C Z, Wang Y L, Nie X W, et al.2019. Evaluation of the effects of sequence length and microsatellite instability on single-guide RNA activity and specificity[J]. International Journal of Biological Sciences, 15(12): 2641-2653. [64] Zhao C Z, Zheng X G, Qu W B, et al.2017. CRISPR-offinder: A CRISPR guide rna design and off-target searching tool for user-defined protospacer adjacent motif[J]. International Journal of Biological Sciences, 13(12): 1470-1478. [65] Zheng Q T, Lin J, Huang J J, et al.2017. Reconstitution of UCP1 using CRISPR/CAS9 in the white adipose tissue of pigs decreases fat deposition and improves thermogenic capacity[J]. Proceedings of the National Academy of Sciences of the USA, 114(45): E9474-E9482. [66] Zou Y L, Li Z Y, Zou Y J, et al.2018. An FBXO40 knockout generated by CRISPR/Cas9 causes muscle hypertrophy in pigs without detectable pathological effects[J]. Biochemical and Biophysical Research Communications, 498(4): 940-945.