Cas13d-Mediated Knockdown of Long Noncoding RNAs

CRISPR-Cas13 systems represent a powerful and programmable platform for RNA-targeted applications, including RNA knockdown. Long noncoding RNA (lncRNA), which lacks an open reading frame and is thereby often refractory to Cas9-mediated indel production, can be successfully knocked down by Cas13 systems. DeepCas13, a deep learning-based model, was developed to predict Cas13d on-target activity for RNA interference with high accuracy. Here, we describe detailed protocols about sgRNA design and experimental procedures for the lentiviral vector-based CRISPR-Cas13 system to knock down individual lncRNAs. Our protocol can be naturally extended to knockdown other types of coding and noncoding RNAs, including messenger RNA (mRNA) circular RNA (circRNA).