129S-H3f3b^tm1.1Mnn/J

Stock number: 021789
Research areas: Cell Biology Research, Research Tools

Description

These H3f3b^c/c mutant mice possess loxP sites flanking a wildtype H3 histone isoform 3 (H3.3) coding sequence and STOP sequence. When bred to mice expressing cre-recombinase, removal of the floxed sequence results in CFP expression. This strain may be useful for studying the role of histone H3.3 during development.

Detailed description

The H3f3b^c/c targeting vector contains, from 5' to 3', a synthetic splice acceptor, a loxP-site, a wildtype H3 histone isoform 3 (H3.3) coding sequence, H3f3b (H3 histone, family 3B) 3' UTR, and an SV40 polyadenylation (polyA) sequence. This was followed by a second loxP site, an ATG start codon, the cerulean variant of cyan fluorescent protein (CFP), and another SV40 polyA sequence. This vector replaced all of exon 2 of the H3f3b gene. H3f3b encodes the histone H3 isoform, H3.3, involved in the regulation of chromatin activity during DNA replication. Homozygous H3f3b^c/c mice are viable and fertile, and express wildtype H3f3b. In the absence of Cre, CFP expression is prevented by the H3.3 coding sequence. When bred to mice that express Cre recombinase, resulting offspring will have the wildtype loxP-flanked H3.3 coding sequence deleted in the cre-expressing tissues, allowing CFP to be expressed.

Development

A targeting vector was designed to contain, from 5' to 3', a synthetic splice acceptor, a loxP site, an ATG start codon, a wildtype H3 histone isoform 3 (H3.3) coding sequence, H3f3b (H3 histone, family 3B) 3' UTR, and an SV40 polyadenylation (polyA) sequence. This was followed by a downstream frt-flanked neomycin resistance (neo) cassette, and a second loxP site, an ATG start codon, the cerulean variant of cyan fluorescent protein (CFP), and another SV40 polyA sequence. This vector replaced all of exon 2 of the H3f3b gene. The construct was electroporated into 129S1/SvImJ-derived A2 embryonic stem (ES) cells. Correctly targeted ES cells were injected into (C57BL/6 J x CBA/CaH) blastocysts and resulting chimeric mice were bred with 129S1;129S4-Gt(ROSA)26Sor^tm1(FLP1)Dym/J transgenic mice to delete the neo cassette. Resulting offspring were crossed to remove the Flp-expressing transgene, resulting in a colony homozygous for the conditional-H3f3b (H3f3b^c/c) allele. These mice were bred to 129S1/SvImJ mice (Stock No. 002448) for at least 5 generations. Upon arrival, mice were bred to 129S1/SvImJ inbred mice for at least one generation to establish the colony.

Allele

Symbol: H3f3b^tm1.1Mnn
Name: targeted mutation 1.1, Jeff Mann
MGI accession ID: MGI:5474376
Generation method: Targeted
Attributes: Conditional ready (e.g. floxed); No functional change
Marker symbol: H3f3b
Marker name: H3 histone, family 3B
Chromosome: 11
Strain of origin: 129S1/SvImJ
Molecular note: A targeting vector was designed to contain, from 5' to 3', a synthetic splice acceptor, a loxP site, an ATG start codon, a wildtype H3 histone isoform 3 (H3.3) coding sequence, H3f3b (H3 histone, family 3B) 3' UTR, and an SV40 polyadenylation (polyA) sequence. This was followed by a downstream frt-flanked neomycin resistance (neo) cassette, and a second loxP site, an ATG start codon, the cerulean variant of cyan fluorescent protein (CFP), and another SV40 polyA sequence. This vector replaced all of exon 2. Flp-mediated recombination removed the neo cassette.