Advances in CRISPR-Cas9 genome editing technology have strengthened the role of zebrafish as a model organism for genetics and developmental biology. These tools have led to a significant increase in the production of loss-of-function mutant zebrafish lines. However, the generation of precisely edited knock-in lines has remained a significant challenge in the field due to the decreased efficiency of homology directed repair (HDR). In this study, we overcame some of these challenges by combining available design tools and synthetic, commercially available CRISPR reagents to generate a knock-in line carrying an in-frame MYC epitope tag at the sox11a locus. Zebrafish Sox11a is a transcription factor with critical roles in organogenesis, neurogenesis, craniofacial, and skeletal development; however, only a few direct molecular targets of Sox11a have been identified. Here, we evaluate the knock-in efficiency of various HDR donor configurations and demonstrate the successful expression and localization of the resulting knock-in allele. Our results provide an efficient, streamlined approach to knock-in experiments in zebrafish, which will enable expansion of downstream experimental applications that have previously been difficult to perform. Moreover, the MYC-Sox11a line we have generated will allow further investigation into the function and direct targets of Sox11a.
|Number of pages||6|
|Journal||Biochemical and Biophysical Research Communications|
|State||Published - Jun 11 2022|
Bibliographical noteFunding Information:
We thank Brandi Bolton, Evelyn Turnbaugh, and Lucas Vieira Francisco for exceptional zebrafish care. We also thank members of the Morris lab for valuable input and technical assistance. This work was supported by grants from the NIH National Eye Institute (R01EY021769, to A.C.M.; F30EY031545, to L.A.K.)
We thank Brandi Bolton, Evelyn Turnbaugh, and Lucas Vieira Francisco for exceptional zebrafish care. We also thank members of the Morris lab for valuable input and technical assistance. This work was supported by grants from the NIH National Eye Institute ( R01EY021769 , to A.C.M.; F30EY031545 , to L.A.K.)
© 2022 Elsevier Inc.
ASJC Scopus subject areas
- Molecular Biology
- Cell Biology