College of Animal Science and Veterinary Medicine, Henan Agricultural University, Zhengzhou, China.
National Research Center for Veterinary Medicine, Luoyang, China.
Transbound Emerg Dis. 2020 Mar;67(2):564-571. doi: 10.1111/tbed.13368. Epub 2019 Sep 30.
Porcine reproductive and respiratory syndrome virus (PRRSV) has varied constantly and circulated in the pig industry worldwide. The prevention and control of porcine reproductive and respiratory syndrome (PRRS) is complicated. A visual, sensitive and specific diagnostic method is advantageous to the control of PRRS. The collateral cleavage activity of LwCas13a is activated to degrade non-targeted RNA, when crRNA of LwCas13a bond to target RNA. The enhanced Cas13a detection is the combination of collateral cleavage activity of LwCas13a and recombinase polymerase amplification (RPA). In this study, the enhanced Cas13a detection for PRRSV was established. The novel method was an isothermal detection at 37°C, and the detection can be used for real-time analysis or visual readout. The detection limit of the enhanced Cas13a detection was 172 copies/μl, and there were no cross-reactions with porcine circovirus 2, porcine parvovirus, classical swine fever virus and pseudorabies virus. The enhanced Cas13a detection can work well in clinical samples. In summary, a visual, sensitive and specific nucleic acid detection method based on CRISPR-Cas13a was developed for PRRSV.
猪繁殖与呼吸综合征病毒（PRRSV）在全球范围内不断变异并在养猪业中传播。猪繁殖与呼吸综合征（PRRS）的防控较为复杂。一种直观、敏感和特异的诊断方法有利于 PRRS 的控制。当 LwCas13a 的 crRNA 与靶 RNA 结合时，LwCas13a 的旁切活性被激活，从而降解非靶向 RNA。增强型 Cas13a 的检测是 LwCas13a 的旁切活性与重组酶聚合酶扩增（RPA）的结合。本研究建立了用于检测 PRRSV 的增强型 Cas13a 检测方法。该新方法在 37°C 下进行等温检测，可用于实时分析或可视化读取。增强型 Cas13a 检测的检测限为 172 拷贝/μl，与猪圆环病毒 2、猪细小病毒、猪瘟病毒和伪狂犬病病毒无交叉反应。该增强型 Cas13a 检测方法可在临床样本中很好地发挥作用。总之，本研究开发了一种基于 CRISPR-Cas13a 的直观、敏感和特异的 PRRSV 核酸检测方法。
