Biomedical Research Core 1: Gene Targeting Core (Core Director: Chris Ward) Core 1 will offer gene targeting for users interested in PKD using both TALENs and CRISPRs, which induce double-stranded DNA breaks that are repaired using the error-prone non-homologous end joining (NHEJ) pathway. NHEJ often results in short deletions or insertions and thus are very efficient and effective in knocking out gene function if targeted to coding regions. The Core will work with users to pin-point the region of the gene to be targeted and the strain to be used. The Core will also supply hypomorphic Pkd1 mice if the investigator wishes to quickly assess the cyst-promoting activity of a heterozygous TALEN or CRISPR-induced null on a mildly cystic `sensitized' background. The Core will also make specific custom alterations (genome edits or knock-in mutations) in the Pkd1 or Pkd2 gene, and will provide services for mutating genes that have been implicated in physically interacting with the PC1/PC2 complex. The Core will also target cell lines (including ES cells) producing matched lines with mutations for physiological cyst forming assays and electrophysiology, and will introduce tandem affinity tags (TAP) tags into the open reading frame of candidate genes to allow the candidate and interacting proteins to be isolated from the cells under physiological conditions, without overexpression. The Gene Targeting Core will work with existing cores at the University of Kansas Medical Center, including the Transgenic and Gene-Targeting Institutional Facility and the Genome Sequencing Facility.
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