Open AccessmRNA processing in mutant zebrafish lines generated by chemical and CRISPR-mediated mutagenesis produces unexpected transcripts that escape nonsense-mediated decay
Author(s) -
Jennifer L. Anderson,
Timothy S. Mulligan,
Meifen Shen,
Hui Wang,
Catherine M. Scahill,
Frederick J. Tan,
Shaojun Du,
Elisabeth M. BuschNentwich,
Steven A. Farber
Publication year - 2017
Publication title -
plos genetics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.587
H-Index - 233
eISSN - 1553-7404
pISSN - 1553-7390
DOI - 10.1371/journal.pgen.1007105
Subject(s) - zebrafish , biology , crispr , genetics , mutagenesis , mutant , nonsense mediated decay , genome editing , nonsense mutation , rna splicing , gene , phenotype , reverse genetics , exon , frameshift mutation , cas9 , mutation , forward genetics , genome , rna , missense mutation
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.