Jilin Provincial Key Laboratory of Animal Embryo Engineering, Institute of Zoonosis, College of Animal Sciences, Jilin University, 130062, Changchun, Jilin Province, People's Republic of China.
Key Laboratory of Jilin Province for Zoonosis Prevention and Control, Institute of Military Veterinary, Academy of Military Medical Sciences, Changchun, Jilin Province, People's Republic of China.
PLoS Pathog. 2018 Dec 13;14(12):e1007193. doi: 10.1371/journal.ppat.1007193. eCollection 2018 Dec.
Classical swine fever (CSF) caused by classical swine fever virus (CSFV) is one of the most detrimental diseases, and leads to significant economic losses in the swine industry. Despite efforts by many government authorities to stamp out the disease from national pig populations, the disease remains widespread. Here, antiviral small hairpin RNAs (shRNAs) were selected and then inserted at the porcine Rosa26 (pRosa26) locus via a CRISPR/Cas9-mediated knock-in strategy. Finally, anti-CSFV transgenic (TG) pigs were produced by somatic nuclear transfer (SCNT). Notably, in vitro and in vivo viral challenge assays further demonstrated that these TG pigs could effectively limit the replication of CSFV and reduce CSFV-associated clinical signs and mortality, and disease resistance could be stably transmitted to the F1-generation. Altogether, our work demonstrated that RNA interference (RNAi) technology combining CRISPR/Cas9 technology offered the possibility to produce TG animal with improved resistance to viral infection. The use of these TG pigs can reduce CSF-related economic losses and this antiviral strategy may be useful for future antiviral research.
经典猪瘟（Classical swine fever，CSF）由经典猪瘟病毒（Classical swine fever virus，CSFV）引起，是一种极具危害性的疾病，给养猪业造成了巨大的经济损失。尽管许多政府部门努力从全国猪群中消灭这种疾病，但它仍然广泛存在。本研究通过 CRISPR/Cas9 介导的基因敲入策略，选择抗病毒小发夹 RNA（small hairpin RNA，shRNA）并插入猪 Rosa26（pRosa26）基因座。最后，通过体细胞核移植（somatic nuclear transfer，SCNT）生产出抗 CSFV 的转基因（transgenic，TG）猪。值得注意的是，体外和体内病毒攻毒试验进一步表明，这些 TG 猪能够有效限制 CSFV 的复制，减轻 CSFV 相关的临床症状和死亡率，并且抗病性可以稳定传递给 F1 代。总之，本研究表明，CRISPR/Cas9 技术与 RNA 干扰（RNA interference，RNAi）技术相结合为生产抗病毒感染能力增强的 TG 动物提供了可能。使用这些 TG 猪可以减少与 CSF 相关的经济损失，这种抗病毒策略可能对未来的抗病毒研究具有重要意义。
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