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mRNA processing in mutant zebrafish lines generated by chemical and CRISPR-mediated mutagenesis produces unexpected transcripts that escape nonsense-mediated decay.

Published version
Peer-reviewed

Type

Article

Change log

Authors

Anderson, Jennifer L  ORCID logo  https://orcid.org/0000-0002-7015-5791
Mulligan, Timothy S  ORCID logo  https://orcid.org/0000-0002-6046-4523
Shen, Meng-Chieh 
Scahill, Catherine M 

Abstract

As model organism-based research shifts from forward to reverse genetics approaches, largely due to the ease of genome editing technology, a low frequency of abnormal phenotypes is being observed in lines with mutations predicted to lead to deleterious effects on the encoded protein. In zebrafish, this low frequency is in part explained by compensation by genes of redundant or similar function, often resulting from the additional round of teleost-specific whole genome duplication within vertebrates. Here we offer additional explanations for the low frequency of mutant phenotypes. We analyzed mRNA processing in seven zebrafish lines with mutations expected to disrupt gene function, generated by CRISPR/Cas9 or ENU mutagenesis methods. Five of the seven lines showed evidence of altered mRNA processing: one through a skipped exon that did not lead to a frame shift, one through nonsense-associated splicing that did not lead to a frame shift, and three through the use of cryptic splice sites. These results highlight the need for a methodical analysis of the mRNA produced in mutant lines before making conclusions or embarking on studies that assume loss of function as a result of a given genomic change. Furthermore, recognition of the types of adaptations that can occur may inform the strategies of mutant generation.

Description

Keywords

Animals, Clustered Regularly Interspaced Short Palindromic Repeats, Codon, Nonsense, Exons, Gene Editing, Gene Expression, Genome, Genomics, Mutagenesis, Mutation, Nonsense Mediated mRNA Decay, RNA Stability, RNA, Messenger, Zebrafish

Journal Title

PLoS Genet

Conference Name

Journal ISSN

1553-7390
1553-7404

Volume Title

13

Publisher

Public Library of Science (PLoS)