Bioscience Methods 2024, Vol.15, No.4, 162-172 http://bioscipublisher.com/index.php/bm 1 68 genes encoding these peptides into the pig genome, researchers can enhance the innate immune response of pigs, providing a built-in defense mechanism against infections. This genetic enhancement is particularly valuable as it offers a proactive approach to pathogen resistance, complementing the passive defense provided by environmental biosafety measures. Other genetic modifications, such as the alteration of immune response genes, further bolster the pig's ability to resist infections. These enhancements ensure that the pigs have an intrinsic capacity to fight off pathogens, reducing the likelihood of disease transmission through transplantable organs. The integration of AMPs and other genetic enhancements into the overall biosafety strategy provides a multi-faceted defense system, significantly improving the resilience of pigs against a wide range of pathogens (Chen et al., 2019). Figure 3 CD163Mut/Mut PAMs are remarkably resistant to HP-PRRSV infection (Adopted from Chen et al., 2019) Image caption: CD163Mut/Mut PAMs are remarkably resistant to HP-PRRSV infection. (A) After infection with the HP-PRRSV strain JXwn06 at the indicated MOIs (0.005, 0.025, 0.1, 0.25, 2.0), culture supernatants were collected at 36 hpi, and viral titers were analyzed by a standard TCID50 assay (left). Cells were collected to measure relative expression of viral RNA by qRT-PCR (right). GAPDH mRNA was used as an endogenous control. (B) After infection with HP-PRRSV strain JXwn06 at an MOI of 0.1, viral titers were measured by TCID50 at the indicated time points (12, 24, 36 and 48 h) (left). Relative expression of viral RNA was analyzed using qRT-PCR (right). GAPDH mRNA was used as an endogenous control. (C) PAMs were infected with JXwn06 at an MOI of 0.1, and 36 h later, levels of PRRSV protein GP5 were analyzed by Western blotting analysis (left). Expression of α-tubulin was shown as a loading control. After 48 h, cells were fixed for detection of PRRSV N protein (Green) by immunofluorescent staining(right). The nuclei (blue) were stained with DAPI. (D) The in vitro infection experiment was carried out with the HP-PRRSV strain WUH3. At the indicated MOIs (0.005, 0.025, 0.1, 0.25, 1.0), viral titers were analyzed by a standard TCID50 assay (left). After infection at an MOI of 0.025, relative expression of viral RNA was analyzed using qRT-PCR at the indicated time points (12, 24, 36, 48, 60 and 72 h). GAPDH mRNA was used as an endogenous control. Data are presented as the mean±SD, n=3. * P<0.05, ** P<0.01, *** P<0.001 (Adopted from Chen et al., 2019)
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