The Pvalb-T2A-dCre-D targeted mutation (also called Pvalb-2A-dCre-D, Pvalb-2A-dCre-Δ or Pvalb-2A-dCre-Δhygro) has a viral 2A oligopeptide (T2A) that mediates ribosomal skipping and a destabilized Cre fusion gene (dCre) inserted in-frame at the 3' end of the parvalbumin (Pvalb) coding region. This is designed to have both endogenous gene and dCre expression directed to Pvalb-expressing cells by the endogenous promoter/enhancer elements.
The ecDHFRR12Y/Y100I domain of dCre leads to proteosomal degradation of the entire fusion protein, resulting in little or no Cre recombinase activity. Administration of the DHFR inhibitor, trimethoprim (TMP), prevents degradation of the dCre fusion gene and results in Cre recombinase activity.
When Pvalb-T2A-dCre-D mice are bred with mice containing loxP-flanked sequences, TMP-stabilized Cre recombination will result in deletion of floxed sequences in the Pvalb-expressing cells of the double mutant offspring.
Specifically, the donating investigator reports that Pvalb-T2A-dCre-D mice have trimethoprim-inducible Cre recombinase expression (in situ hybridization) that is specific and efficient in scattered cells throughout the cortex, as well as restricted populations in the cerebellum, medulla, pons, pallidum, and thalamus.
In the absence of trimethoprim, these same regions exhibit significantly reduced Cre recombinase activity (highest expression in particular cells of the cerebellum and in the thalamic reticular nucleus). Heterozygous mice are viable and fertile with no gross physical or behavioral abnormalities. The donating investigator did not examine dCre activity in tissues other than brain, and did not attempt to generate homozygous mice to date (July 2013).
For characterization information, see images at the Allen Institute for Brain Science website (Pvalb-T2A-dCre images).
The dCre fusion gene (also called destabilized Cre, hDHFR/Cre or ecDHFRR12Y/Y100I/Cre) is Cre recombinase fused at its N terminus to the first 159 amino acids of the Escherichia coli K-12 strain chromosomal dihydrofolate reductase gene (DHFR or folA) harboring the G67S mutation and modified to also include the R12Y/Y100I destabilizing domain mutations. The ecDHFRR12Y/Y100I domain of dCre leads to proteosomal degradation of the entire fusion protein, resulting in little or no Cre recombinase activity. The donating investigator reports that administration of the high affinity DHFR inhibitor trimethoprim (TMP; at a concentration of 0.25-0.30 mg/g body weight) prevents degradation of the dCre fusion gene, resulting in Cre recombinase activity.
