Researchers evaluated the use of a dual clustered regularly interspaced short palindromic repeats (CRISPR)-Cas3 system.
By Pooja Toshniwal PahariaAug 29 2023Reviewed by Lily Ramsey, LLM In a recent study published in Stem Cell Reports, researchers evaluated the use of a dual clustered regularly interspaced short palindromic repeats -Cas3 system for inducing multi-exon skipping among Duchenne muscular dystrophy -patient-derived induced pluripotent stem cells .
About the study The present study assessed the CRISPR-Cas3 system as a genome editing tool for DMD patients. A pair of crRNAs inwardly sandwiching the target genomic region was used for the analysis. The dual Cas3 approach was enriched using the SSA reporter vector system adapted for the dual Cas3. Immunostaining and Western blot analyses were performed to assess dystrophin protein recovery, and reverse transcription PCR was used to confirm MES induction at the messenger ribonucleic acid level.
Genome editing using the dual CRISPR-Cas3 system was performed in DMD patient-obtained induced pluripotent stem cell lines FF12020, CiRA00458, and CiRA00646. No significant off-target deletions were found at the putative crRNA binding sites. The inward-type dual CRISPR-Cas3 method effectively caused effective deletions of up to 344 kb among HEK293T cells and DMD patient-obtained iPSCs.
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