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Characterization of Poldip2 knockout mice: Avoiding incorrect gene targeting
Bernard Lassègue1, Sandeep Kumar2, Rohan Mandavilli1
1Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA, United States of America.
Plos One
|December 20, 2021
Summary
Researchers developed a new conditional Poldip2 mouse model to study the protein's function. This model overcomes limitations of previous studies, revealing POLDIP2 inactivation impacts multiple cellular processes.
Area of Science:
- Molecular Biology
- Genetics
- Genomics
Background:
- POLDIP2 is a multifunctional protein with incompletely understood roles.
- Previous mouse models had limitations including lethality and unintended gene downregulation.
- A new conditional floxed Poldip2 mouse model was developed to address these issues.
Purpose of the Study:
- To develop and validate a new conditional floxed Poldip2 mouse model.
- To generate a constitutive Poldip2 knockout mouse line.
- To investigate the physiological and molecular effects of Poldip2 ablation in vasculature.
Main Methods:
- Generation of a conditional floxed Poldip2 mouse model.
- Detection of unexpected genomic duplications using advanced molecular techniques.
- Creation of a constitutive knockout line via Cre-mediated recombination.
- RNA-sequencing (RNA-seq) and quantitative real-time PCR (RT-qPCR) on knockout arteries.
Main Results:
- An initial floxed mouse line exhibited an undetected 305 kb duplication around Poldip2.
- A corrected floxed Poldip2 model allowed generation of viable homozygous knockout mice.
- Poldip2 ablation in arteries affects multiple cellular processes, as shown by transcriptomic analysis.
Conclusions:
- The new conditional Poldip2 model overcomes limitations of previous studies.
- Genome editing methods require careful validation to detect unexpected mutations like duplications.
- POLDIP2 plays a significant role in vascular cellular processes, warranting further investigation.
