Xiao, A., Wang, Z., Hu, Y., Wu, Y., Luo, Z., Yang, Z., Zu, Y., Li, W., Huang, P., Tong, X., Zhu, Z., Lin, S., and Zhang, B. (2013) Chromosomal deletions and inversions mediated by TALENs and CRISPR/Cas in zebrafish. Nucleic acids research. 41(14):e141.
Customized TALENs and Cas9/gRNAs have been used for targeted mutagenesis in zebrafish to induce indels into protein-coding
genes. However, indels are usually not sufficient to disrupt the function of non-coding genes, gene clusters or regulatory
sequences, whereas large genomic deletions or inversions are more desirable for this purpose. By injecting two pairs of TALEN
mRNAs or two gRNAs together with Cas9 mRNA targeting distal DNA sites of the same chromosome, we obtained predictable genomic
deletions or inversions with sizes ranging from several hundred bases to nearly 1 Mb. We have successfully achieved this type
of modifications for 11 chromosomal loci by TALENs and 2 by Cas9/gRNAs with different combinations of gRNA pairs, including
clusters of miRNA and protein-coding genes. Seven of eight TALEN-targeted lines transmitted the deletions and one transmitted
the inversion through germ line. Our findings indicate that both TALENs and Cas9/gRNAs can be used as an efficient tool to
engineer genomes to achieve large deletions or inversions, including fragments covering multiple genes and non-coding sequences.
To facilitate the analyses and application of existing ZFN, TALEN and CRISPR/Cas data, we have updated our EENdb database
to provide a chromosomal view of all reported engineered endonucleases targeting human and zebrafish genomes.