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Abstract: TH-PO314

Generation of a ATP6V1G3-Cre Mice With High Kidney Intercalated Cell Specificity

Session Information

Category: Fluid‚ Electrolyte‚ and Acid-Base Disorders

  • 1001 Fluid‚ Electrolyte‚ and Acid-Base Disorders: Basic


  • Saxena, Vijay, Indiana University School of Medicine, Indianapolis, Indiana, United States
  • Arregui, Samuel, Indiana University School of Medicine, Indianapolis, Indiana, United States
  • Hains, David S., Indiana University School of Medicine, Indianapolis, Indiana, United States
  • Schwaderer, Andrew L., Indiana University School of Medicine, Indianapolis, Indiana, United States

The kidney collecting duct is composed of principal cells (PCs) and intercalated cells (ICs). Principal cells maintain sodium and water balance while intercalated cells maintain acid-base homeostasis. Recent studies have shown that ICs are the critical cells to defend against uropathogen invasion. For focused studies of IC cells, a novel Atp6v1b1-Cre (B1-Cre) mice (generated by Dr. Raoul Nelson group) were crossed with Tdtomato-flox mice to enrich ICs from kidney. B1-Cre mice strain is very useful for enriching ICs from kidney, we noticed that they also labeled kidney resident CD45+ immune cells hence limiting their use for enrichment and gene expression studies. Additionally, Atp6v1b1 is expressed in a range of tissues including the lung, brain, pancreas, skin and gastrointestinal tract. Here we report the generation of Atp6v1g3-Cre mice using CRISPR/CAS technology which labels kidney ICs without any expression on kidney resident immune cells and minimal extra kidney expression.


To generate the mouse, a CRISPR targeting site, Atp6v1g3 gRNA 76/59 in intron 1 nearby the 3’ end of exon 1 with acceptable predicted efficiency and specificity was designed. A T2A-cre-PolyA cassette was inserted into exon1 by CRISPR/Homology directed repair (HDR) strategy. The resulting transcript under the control of endogenous Atp6v1g3 promoter express a short peptide (27aa of Atp6v1g3 N terminal and 17aa of T2A) and the Cre recombinase. CRISPR/cas9 gRNA was injected to the zygotes of pronuclei of fertilized eggs and F0 progeny was generated. F0 progeny mice were genotyped by Real-Time PCR (RT-PCR) for G3-Cre expression and bred to wild type C57BL/6 mice to maintain colony of mice.


Atp6v1g3-Cre were crossed with Tdtomatoflox/flox mice to generate Atp6v1g3-Cre+/Tdtomatoflox/- mice. ICs were flowsorted and their relative enrichment was confirmed by RT-PCR. Tdtomato expression was analyzed on Flowcytometer and immunofluorescence which revealed expression on ICs but not on CD45+ immune cells along with minimal extra kidney expression in various tissues as examined by immunofluorescence and RT-PCR.


The Atp6v1g3-Cre mouse will be a useful tool for kidney IC specific studies.


  • NIDDK Support