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Construction of PK-15 cell line withLTA4H gene knockout and its impact on the replication of foot-and-mouth disease virus
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Jiali WANG1, 2, 3, Xiuying PU1, Fan YANG2, 3, Wenhua SHAO2, 3, Mengyao HUANG2, 3, Weijun CAO2, 3, Haixue ZHENG1, 2, 3, *, Wei ZHANG2, 3, *
Acta Microbiologica Sinica | 2024, 64(3) : 733 - 744
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Acta Microbiologica Sinica | 2024, 64(3): 733-744
• Research Articles •
Construction of PK-15 cell line withLTA4H gene knockout and its impact on the replication of foot-and-mouth disease virus
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Jiali WANG1, 2, 3, Xiuying PU1, Fan YANG2, 3, Wenhua SHAO2, 3, Mengyao HUANG2, 3, Weijun CAO2, 3, Haixue ZHENG1, 2, 3, *, Wei ZHANG2, 3, *
Affiliations
  • 1 School of Life Science and Engineering, Lanzhou University of Technology, Lanzhou 730050, Gansu, China
  • 2 State Key Laboratory of Animal Disease Prevention and Control, Lanzhou Veterinary Institute, Chinese Academy of Agricultural Sciences/College of Animal Medicine and Biosafety, Lanzhou University, Lanzhou 730000, Gansu, China
  • 3 Gansu Provincial Research Center of Pathogen Biology, Lanzhou 730046, Gansu, China
Published: 2024-03-04 doi: 10.13343/j.cnki.wsxb.20230500
Outline
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[Objective] ALTA4H-deleted porcine kidney cell line (PK-15) was constructed by CRISPR/Cas9 to study the effect ofLTA4H on the replication of foot-and-mouth disease virus (FMDV), with a view to providing a theoretical basis for revealing the functions ofLTA4H and the mechanism of regulating virus replication. [Methods] Two small guide RNAs (sgRNAs) targeting porcineLTA4H were designed and integrated into the vector pX459-puro-MCS, respectively. The recombinant plasmid was transfected into PK-15 cells, which were then cultured in the medium supplemented with puromycin, and the monoclonal cells were selected by limiting dilution method. Western blotting and sequencing were performed to detectLTA4H knockout, andLTA4H-deleted cells were obtained. Furthermore, Western blotting, RT-qPCR, and virus titer assay were employed to examine FMDV replication and protein expression afterLTA4H knockout. [Results] Compared with the wild type cells, theLTA4H-deleted PK-15 cells exhibited inhibited FMDV replication. [Conclusion] This study successfully constructed a PK-15 cell line with LTA4H gene knockout, demonstrating that LTA4H can promote the replication of FMDV, and the results provided a theoretical basis for the subsequent research on the function ofLTA4H.

CRISPR/Cas9  /  leukotriene A4 hydrolase  /  cell lines  /  foot-and-mouth disease virus
Jiali WANG, Xiuying PU, Fan YANG, Wenhua SHAO, Mengyao HUANG, Weijun CAO, Haixue ZHENG, Wei ZHANG. Construction of PK-15 cell line withLTA4H gene knockout and its impact on the replication of foot-and-mouth disease virus[J]. Acta Microbiologica Sinica, 2024 , 64 (3) : 733 -744 . DOI: 10.13343/j.cnki.wsxb.20230500
Funding
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  • National Natural Science Foundation of China(32102639)
  • National Natural Science Foundation of China(32072831)
  • National Key Research and Development Program of China(2021YFD1800300)
  • Distinguished Young Scholars Fund of Gansu Province(21JR7RA026)
  • Basic Research Fund of the Central University of Lanzhou University(lzujbky-2022-ey20)
  • Open Competition Program of Top Ten Critical Priorities of Agricultural Science and Technology Innovation for the 14th Five-Year Plan of Guangdong Province(2023SDZG02)
  • National Technical System for Pig Industry(CARS-35)
Appendix
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Year 2024 volume 64 Issue 3
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85
35
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Article Info
doi: 10.13343/j.cnki.wsxb.20230500
  • Receive Date：2023-07-29
  • Online Date：2026-03-19
  • Published：2024-03-04
Article Data
Affiliations
History
  • Received：2023-07-29
  • Accepted：2023-12-13
Funding
National Natural Science Foundation of China(32102639)
National Natural Science Foundation of China(32072831)
National Key Research and Development Program of China(2021YFD1800300)
Distinguished Young Scholars Fund of Gansu Province(21JR7RA026)
Basic Research Fund of the Central University of Lanzhou University(lzujbky-2022-ey20)
Open Competition Program of Top Ten Critical Priorities of Agricultural Science and Technology Innovation for the 14th Five-Year Plan of Guangdong Province(2023SDZG02)
National Technical System for Pig Industry(CARS-35)
Affiliations
    1 School of Life Science and Engineering, Lanzhou University of Technology, Lanzhou 730050, Gansu, China
    2 State Key Laboratory of Animal Disease Prevention and Control, Lanzhou Veterinary Institute, Chinese Academy of Agricultural Sciences/College of Animal Medicine and Biosafety, Lanzhou University, Lanzhou 730000, Gansu, China
    3 Gansu Provincial Research Center of Pathogen Biology, Lanzhou 730046, Gansu, China

Corresponding:

*ZHENG Haixue, E-mail:;
ZHANG Wei, E-mail:
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表12种不同金属材料的力学参数
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Family		 属数
Number of
genus		 种数
Number of
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Percentage of
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鹅膏菌科Amanitaceae 2 11 5.26 鹅膏菌属 Amanita 10 4.78
小菇科 Mycenaceae 2 12 5.74 丝盖伞属 Inocybe 5 2.39
多孔菌科 Polyporaceae 8 14 6.70 蜡蘑属 Laccaria 5 2.39
红菇科 Russulaceae 3 23 11.00 小皮伞属 Marasmius 6 2.87
小菇属 Mycena 11 5.26
光柄菇属 Pluteus 5 2.39
红菇属 Russula 17 8.13
栓菌属 Trametes 5 2.39
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