Multi-resistance strategy for viral diseases and short hairpin RNA verification method in pigs.

OBJECTIVE

Foot and mouth disease (FMD) and porcine reproductive and respiratory syndrome (PRRS) are major diseases that interrupt porcine production. Because they are viral diseases, vaccinations are of only limited effectiveness in preventing outbreaks. To establish an alternative multi-resistant strategy against FMD virus (FMDV) and PRRS virus (PRRSV), the present study introduced two genetic modification techniques to porcine cells.

METHODS

First, cluster of differentiation 163 (CD163), the PRRSV viral receptor, was edited with the clustered regularly interspaced short palindromic repeats-CRISPR-associated protein 9 technique. The gene sequences of edited cells and control cells differed. Second, short hairpin RNA (shRNAs) were integrated into the cells. The shRNAs, targeting the gene of FMDV and the open reading frame 7 () gene of PRRSV, were transferred into fibroblasts. We also developed an shRNA verification method with a target gene expression vector.

RESULTS

shRNA activity was confirmed with vectors that expressed the and genes in the cells. Cells containing shRNAs showed lower transcript levels than cells with only the expression vectors. The shRNAs were integrated into CD163-edited cells to combine the two techniques, and the viral genes were suppressed in these cells.

CONCLUSION

We established a multi-resistant strategy against viral diseases and an shRNA verification method.