Unexpected extra exon skipping in the DYSF gene during restoring the reading frame by CRISPR/Cas9

In this study, we evaluated the possibility of permanent skipping of exons 3-4, and 26-27 which deletion does not violate the reading frame and allows to eliminate truncated variants within exons. Editing was performed with simultaneous transfection of two sgRNA- and sa/spCas9-containing plasmids on HEK293T cell cultures and healthy donor myoblasts. Skipping of exons 3-4 was performed by destroying the splicing acceptor sites, and exons 26-27 by cuts in the flanking exons with the corresponding deletion in the DNA. Some unexpected results were obtained, when exons 26-27 were skipped, exon 30 was also absent in the transcript, although it is not alternatively spliced and is normally present in all transcripts. This event indicates that DNA changes near splicing sites can affect adjacent exons and the whole gene. However, this fact requires further study.PMID:37944631 | DOI:10.1016/j.biosystems.2023.105072

Source: Biosystems - November 9, 2023 Category: Biotechnology Authors: Olga Levchenko Irina Panchuk Konstantin Kochergin-Nikitsky Irina Petrova Sabina Nagieva Maxim Pilkin Ivan Yakovlev Svetlana Smirnikhina Roman Deev Alexander Lavrov Source Type: research