B6.Cg-Dlg4^tm1.2Hnz/J

Stock number: 029242
Product name: PSD-95-CreNABLED
Research areas: Neurobiology Research, Research Tools

Description

This strain expresses functional mVenus-labeled PSD95 (DLG4) synaptic protein.

Detailed description

Dlg4 (discs, large homolog 4 (Drosophila); also called PSD95) is a postsynaptic protein that plays a pivotal role in synaptic function and plasticity.

PSD-95-CreNABLED mice, derived from a cross of PSD-95-ENABLED mice (see Stock No. 026092) and germline HPRT-Cre mice (see Stock No. 004302), express functional mVenus-labeled DLG4 synaptic protein.

Strong mVenus fluorescence can be detected throughout many brain regions, including hippocampus, neocortex, striatum and thalamus. Relatively little fluorescence is observed in the cerebellum, consistent with the current knowledge that cerebellar Purkinje neurons do not express DLG4. At higher magnification under a two-photon microscope, dense green fluorescent puncta are found throughout the neuropil of the neocortex, striatum and hippocampus.

Expression levels are not reduced and synaptic transmission and long-term potentiation are unaltered. The resultant protein functionally replaces that of the wildtype. Label is present in most dendritic spines and exhibits minimal baseline trafficking. This strain enables the visualization and unambiguous examination of shaft excitatory synapses, such as those in aspiny interneurons, in vivo.

Development

The last two exons of the gene (exons 19 and 20), including the 3' UTR, were duplicated and inserted downstream of the polyadenylation signal and a neomycin selection cassette flanked by FRT sites. Two loxP sites were inserted to flank the native exons 19 and 20 as well as a neomycin stop cassette. The sequence for mVenus was fused in frame to the end of the coding sequence in exon 20. The mutation was created through homologous recombination in G1 129S6 x C57BL/6J F1-derived embryonic stem (ES) cells. The FRT-flanked neomycin cassette was subsequently excised through crosses with a germline FLP strain (see Stock No. 003946) on a 129S1/Sv genetic background. Animals were crossed with germline-specific Hprt-Cre mice (see Stock No. 004302) to excise the floxed exons 19 and 20 and enable expression of mVenus-labeled PSD95 (DLG4) synaptic protein. This strain was backcrossed to C57BL/6NCrl for 5 generations by the donating lab.

Allele

Symbol: Dlg4^tm1.2Hnz
Name: targeted mutation 1.2, Haining Zhong
MGI accession ID: MGI:5770125
Generation method: Targeted
Attributes: Reporter
Marker symbol: Dlg4
Marker name: discs large MAGUK scaffold protein 4
Chromosome: 11
Strain of origin: (129S6/SvEvTac x C57BL/6J)F1
Expressed genes: YFP,Yellow Fluorescent Protein,jellyfish
Site of expression: Strong mVenus fluorescence can be detected throughout many brain regions, including hippocampus, neocortex, striatum and thalamus.
Molecular note: The last two exons of the gene (exons 19 and 20), including the 3' UTR, were duplicated and inserted downstream of the polyadenylation signal and a neomycin selection cassette flanked by FRT sites. Two loxP sites were inserted to flank the native exons 19 and 20 as well as a neomycin stop cassette. The sequence for mVenus was fused in-frame to the end of the coding sequence in exon 20. The original mutation was generated by homologous recombination in embryonic stem (ES) cells. The FRT-flanked neomycin cassette was subsequently excised via a cross to a germline FLP-expressing strain, yielding Dlg4^tm1.1Hnz. Mice bearing this allele were bred with germline Cre-expressing mice to excise the floxed exons 19 and 20 and enable expression of mVenus-labeled, functional PSD95 (DLG4) synaptic protein.

DLG4 expression levels are slightly lower in mutant than in wild-type brains. Strong mVenus fluorescence can be detected throughout many brain regions, including hippocampus, neocortex, striatum and thalamus. Relatively little fluorescence is observed in the cerebellum. At higher magnification under a two-photon microscope, dense green fluorescent puncta are found throughout the neuropil of the neocortex, striatum and hippocampus. Label is present in most dendritic spines and exhibits minimal baseline trafficking. This mutation enables the in vivo visualization and unambiguous identification of excitatory shaft synapses in aspiny interneurons.
General note: ES cell line = G1 (129S6/SvEvTac x C57BL/6J)F1.