Cdk6^T2A-drePBD* knock-in mouse

Background information on Cdk6 expression in the brain.

Cdk6 expression in single cell RNA sequencing clusters.

The Cdk6^T2A-td-sfGFP allele targeting vector plasmid was deposited at Addgene for sharing. The NGS sequence is available at https://www.addgene.org/134321/sequences/. One can verify that the homology arms map to Cdk6 using NCBI BLAST.

Cdk6 expression characterization in Cdk6^T2A-td-sfGFP mice.

The Cdk6^T2A-drePBD* allele targeting vector plasmid was deposited at Addgene for sharing. The NGS sequence is available at https://www.addgene.org/141452/sequences/. One can verify that the homology arms map to Cdk6 using NCBI BLAST.

The Cdk6^T2A-drePBD* allele mouse production.

Using the Hipp11-dre reporter (see here), reporter recombination could be induced with RU-486. KI-67- ASCL1- GFP+ cells with neuroblast morphology were revealed. (I couldn't try the anti-DCX antibodies because I didn't have the necessary DyLight405-conjugated goat anti-guinea pig or goat anti-rabbit secondary.) I suppose the induction could have initially labeled C type transit amplifying cells (TAC's). These C type TAC's must have been late in the neurogenic lineage, which in 3 days had differentiated into presumptive neuroblasts. These were consistent with the scRNA-seq dataset analysis, wherein the Cdk6 expression level peaked in the late C type TAC's.

Thus, the Cdk6^T2A-drePBD* allele seemed to work as it should. Nevertheless, the RU-486 inducer unfortunately did not solubilize at higher concentration in corn oil/ethanol vehicle, limiting the inducer dosage. In my experience with the tamoxifen inductions, injecting more frequently at lower dose didn't necessarily induce more recombination. So, I didn't spend much more time injecting more RU-486 with this mouse line. I could have but did not optimize the vehicle to see if I can make a higher concentration formulation because the RU-486 was not cheap (https://www.sigmaaldrich.com/US/en/product/sigma/m8046?context=product) and I wasn't interested in C cells.

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Cdk6 mRNA is also present in activated stem cells, at a lower level perhaps (see avg_expression_genes.txt). The activated stem cells have larger nuclei, as we and others have observed. In my whole mount immunos of Cdk6^{T2A-td-sfGFP/+} mouse brains, some CDK6-T2A-td-sfGFP+ cells with large nucleus were observed. However, activated stem cells were apparently not labeled by the Cdk6^{T2A-drePBD*/+} mouse.

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The Cdk6^T2A-td-sfGFP mice will be available from Jax (# 036458).

We refer interested investigators to Jax to obtain this mouse line.