Additional file 11: of Streamlined computational pipeline for genetic background characterization of genetically engineered mice based on next generation sequencing data

Validation of the murine Sall2 gene deletion by CRISPR-Cas9. A) Representative Western blot for SALL2 and ACTIN in control and Sall2-silenced cells by CRISPR (mSall2 CRISPR) done in Sall2 WT iMEFs. B) We designed a double CRISPR cut to delete a segment of the Sall2 gene. The two CRISPRs (denoted as gRNA one and two) targeted the largest exon of the murine Sall2 gene (exon 2). C). iMEF cells were electroporated with Control CRISPR plasmid or the two mSall2 CRISPR plasmids, and fluorescent cells were enriched by flow-cell cytometry (top 5% of fluorescent cells). We identified the desired deletion from the genomic DNA of a pool of iMEF cells and targeted it with the double CRISPR strategy (amplicon at 500 base pairs in mSall2 lane, denoted with a black arrow). D) Alignment from the Sanger sequencing results of the gel-purified amplicon from (C), depicting the genomic deletion of the Sall2 gene (chromosomal position 52,314,428–52,315,642 on the mm10 build). We highlighted the codifying sequences of the exon two of murine Sall2 gene in yellow. (TIF 808 kb