Ai65F mice (also called Ai65F(RCF-tdT), RCF-tdTomato or FSF-tdTomato) are a fluorescent FLP reporter strain. Ai65F is a Rosa-CAG-FrtSTOPFrt-tdTomato conditional allele designed with a frt-flanked STOP cassette preventing transcription of the tdTomato fluorescent protein. After removal of the frt-flanked STOP cassette by FLP-recombination, tdTomato fluorescence is expected in cells/tissues expressing FLP recombinase.
Of note, Ai65F is a derivative of the dual Cre:FLP fluorescent reporter strain Ai65(RCFL-tdT)-D - which is also available from The Jackson Laboratory as Stock No. 021875.
Hongkui Zeng, Allen Institute for Brain Science
Ai65F mice (also called Ai65F(RCF-tdT), RCF-tdTomato or FSF-tdTomato) are a fluorescent FLP reporter strain. Ai65F is a Rosa-CAG-FrtSTOPFrt-tdTomato conditional allele designed with a frt-flanked STOP cassette preventing transcription of the tdTomato fluorescent protein. Although under control of the endogenous Gt(ROSA)26Sor promoter/enhancer regions and the CAG hybrid promoter, widespread expression of tdTomato is prevented by the STOP cassette. Upon exposure to FLP recombinase, the STOP cassette is removed - resulting in tdTomato fluorescence.
For example, Ai65F allows characterization of FLP-expressing mouse lines [Daigle et al. 2018 Cell 174:465 (PMID:30007418)].
For additional characterization information, see images at the Allen Institute for Brain Science website (Ai65F images).
Prior to introduction of FLP recombinase, mice heterozygous for Ai65F are viable and fertile with no reported gross physical or behavioral abnormalities. To date (December 2018), it has not been attempted to make this strain homozygous.
Ai65F mice (Stock No. 032864 ; also called Ai65F(RCF-tdT), RCF-tdTomato or FSF-tdTomato) were generated in 2018 by breeding Ai65D mice (Stock No. 021875) with a C57BL/6J-congenic germline Cre-deleter strain to remove the loxP-flanked STOP cassette, and then additional matings to C57BL/6J inbred mice removed the Cre allele. Specific details are below.
The Ai65(RCFL-tdT)Δneo allele (also called Ai65(RCFL-tdT)-D, Ai65Δneo, or Ai65D) was created in the laboratory of Dr. Hongkui Zeng (Allen Institute for Brain Science) to have the following inserted into the Gt(ROSA)26Sor locus on chromosome 6 (from 5' to 3'): a CMV-IE enhancer/chicken beta-actin/rabbit beta-globin hybrid promoter (from pCAGGS), a frt-flanked STOP cassette (with a short open reading frame, followed by translational stops in all three reading frames, a synthetic polyA sequence, a polyA sequence from the human growth hormone gene, and a polyA sequence from the Herpes Simplex Virus TK gene), a loxP-flanked STOP cassette (with stop codons in all three reading frames linked to three repeats of the SV40 polyA signal), a sequence encoding the tdTomato fluorescent protein (a non-oligomerizing DsRed fluorescent protein variant with a 12 residue linker fusing two copies of the protein [tandem dimer]), a woodchuck hepatitis virus post-transcriptional regulatory element (WPRE; to enhance the mRNA transcript stability), a bovine growth hormone polyA signal and an AttL site. In 2013, Ai65D mice were sent to The Jackson Laboratory Repository as Stock No. 021875. Upon arrival, some were bred at least two additional generations to C57BL/6J inbred mice (Stock No. 000664).
In 2018, Ai65D mice (backcrossed a total of at least four generations onto C57BL/6J) were selected and bred to C57BL/6J-congenic Sox2-Cre mice (Stock No. 008454) for germline removal of the loxP-flanked STOP cassette. The resulting mice with Ai65F allele (Rosa::CAG::frt-STOP-frt::loxP::tdTomato::WPRE::bGHpA::AttL) were bred at least one additional generation to C57BL/6J inbred mice (and the Sox2-Cre allele was removed). The C57BL/6J-congenic Ai65F colony is Stock No. 032864.
|Allele Name||targeted mutation 65.2, Hongkui Zeng|
|Allele Type||Targeted (Conditional ready (e.g. floxed), Reporter)|
|Allele Synonym(s)||Gt(ROSA)26Sortm65.2(CAG-tdTomato)Hze; targeted mutation 65.2, Hongkui Zeng|
|Gene Symbol and Name||Gt(ROSA)26Sor, gene trap ROSA 26, Philippe Soriano|
|Gene Synonym(s)||beta geo; ROSA26; R26; Gtrgeo26; Gtrosa26; Gtrgeo26; Gtrosa26; AV258896; expressed sequence AV258896; gene trap ROSA 26; gene trap ROSA b-geo 26; SETD5-AS1; Thumpd3as1|
|Strain of Origin||(129S6/SvEvTac x C57BL/6NCrl)F1|
|Molecular Note||The targeting construct contains (from 5' to 3'): a CMV-IE enhancer/chicken beta-actin/rabbit beta-globin hybrid promoter (from pCAGGS), a frt-flanked STOP cassette (with a short open reading frame, followed by translational stops in all three reading frames, a synthetic polyA sequence, a polyA sequence from the human g rowth hormone gene, and a polyA sequence from the Herpes Simplex Virus TK gene), a loxP-flanked STOP cassette (with stop codons in all three reading frames linked to three repeats of the SV40 polyA signal), a sequence encoding the tdTomato fluorescent protein (a non-oligomerizing DsRed fluorescent protein variant with a 12 residue linker fusing two copies of the protein [tandem dimer]), a woodchuck hepatitis virus post-transcriptional regulatory element (WPRE; to enhance the mRNA transcript stability), a bovine growth hormone polyA signal and an AttL site.This entire targeting vector was inserted between exons 1 and 2. PhiC31-mediated recombination removed the AttB/AttP-flanked PGK-Neo-polyA cassette and replace it with the recombined AttB/AttP site (AttL). This entire targeting vector was inserted between exons 1 and 2. PhiC31-mediated recombination removed the AttB/AttP-flanked PGK-Neo-polyA cassette and replaced it with the recombined AttB/AttP site (AttL). Cre-mediated recombination removed the floxed STOP.|
When maintaining a live colony, heterozygous mice may be bred together, to wildtype mice from the colony or to C57BL/6J inbred mice (Stock No. 000664). To date (December 2018), it has not been attempted to make this strain homozygous.
When using the Ai65F , Ai65F(RCF-tdT) , RCF-tdTomato , FSF-tdTomato mouse strain in a publication, please cite the originating article(s) and include JAX stock #032864 in your Materials and Methods section.
We will fulfill your order by providing at least two carriers for each strain ordered. The total number, sex, and genotypes provided will vary, although typically 8 or more animals are provided. Please check genotypes which will be recovered. While the genotypes of all animals produced will be communicated to you prior to scheduling shipment, the genotypes of animals provided may not reflect the mating scheme and genotypes described in the strain description. Animals are typically ready to ship in 11-14 weeks. If a second recovery is required to produce the minimum number of animals, then delivery time would increase to approximately 25 weeks. If we fail to produce animals of the correct genotype, you will not be charged. We cannot guarantee the reproductive success of mice shipped to your facility. If the mice are lost after the first three days (post-arrival) or do not produce progeny at your facility, a new order and fee will be necessary.
Cryorecovery to establish a Dedicated Supply for greater quantities of mice. Mice recovered can be used to establish a dedicated colony to contractually supply you mice according to your requirements. Price by quotation.
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