Strain Name:
B6J.Cg-Gt(ROSA)26Sorem1(CAG-cas9)Jdd/Mmmh
Stock Number:
068058-MU
Citation ID:
RRID:MMRRC_068058-MU
Other Names:
LSL-Rosa26 Cag Cas9 MR04

Strain Information

Gt(ROSA)26Sorem1(CAG-cas9)Jdd
Name: gene trap ROSA 26, Philippe Soriano; endonuclease-mediated mutation 1, Joseph Dougherty
Type: Allele
Species: Multi-species
Chromosome: 6
CAG
Name: CMV early enhancer/chicken beta actin promoter/rabbit beta globin splice acceptor
Type: DNA Segment
Species: Multi-species
Chromosome:
Gt(ROSA)26Sor
Name: gene trap ROSA 26, Philippe Soriano
Synonyms: beta geo, R26, Gtrosa26, Thumpd3as1, Gtrgeo26, ROSA26
Type: Gene
Species: Mus musculus (mouse)
Chromosome: 6
NCBI: 14910
cas9
Name: CRISPR associated protein 9
Type: Gene
Species: Streptococcus pyogenes (bacterium)
Chromosome:
Genetic Alterations
This is a Cre-dependent, Cas9-expressing mouse line with CAG enhancer, Lox-Stop-Lox cassette, and 3xFlag-NLS-Cas9-NLS cloned downstream of the stop cassette, inserted into the Rosa26 locus.
ES Cell Line
Not applicable
Phenotype
None/Normal/Wild-type.

Conditional phenotype: It will conditionally express Cas9

Strain Development
To generate a Cre-dependent Cas9-expressing mouse line, the donor engineered Cas9 into a targeting vector containing Rosa26 homology arms (~1 kb, PCR’s from genome), flanking a CAG enhancer, Lox-Stop-Lox cassette (adapted from the Allen AI9 constructs, RRID:Addgene_22799), with 3xFlag-NLS-Cas9-NLS cloned downstream of the stop cassette. The Flag tag is in-frame to facilitate confirmation of protein expression. The targeting vector was sequence-confirmed then purified for injection into (C57BL/6 x CBA)F1 hybrid oocytes in conjunction with mRNA coding for a pair of custom TALENs designed to target the mouse ROSA26 locus between the homology arms using the ZiFit targeter and ROSA26 TALENs binding sites as follows: (1) 5’ ROSA26 TALEN binding site: 5’-tccctcgtgatctgcaactcc-3’, and (2) 3’ ROSA26 TALEN binding site: 5’-gggcgggagtcttctgggca-3’.

Founders may have been mosaic with multiple mutant alleles and thus were bred to WT to generate heterozygous F1 offspring. Analysis of F1 offspring via long-range PCR confirmed germline transmission of the correctly targeted allele. Such progeny of founder MR04 were bred to a ubiquitous germline Actin-Cre mouse line to confirm Cre-dependence of Cas9 expression. Subsequent generations of Cre- mice were backcrossed to C57BL/6J wild-type mice.
Suggested Control Mice
C57BL6/J
MMRRC Genetic QC Summary
The MMRRC Centers have developed a genetic QC pipeline using MiniMUGA array genotyping to provide additional information on strain backgrounds for MMRRC congenic and inbred strains. For more information on when data may be available, or to request genotyping for a strain of interest, please contact mmrrc@missouri.edu. Older strains may not have this information.
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Donor
Joseph Dougherty, Ph.D., Washington University School of Medicine in St. Louis.
Primary Reference

https://www.biorxiv.org/content/10.1101/2020.03.13.991224v1 and with very minor revisions at Nat Comm.

Colony and Husbandry Information

Colony Surveillance Program and Current Health Reports

Mice recovered from a cryo-archive will have health surveillance performed on recipient females. Health reports will be provided prior to shipment. If you require additional health status information, please email mmrrc@missouri.edu.
Coat Color
Black
Eye
Black
MMRRC Breeding System
Backcross
Generation
N5+ (C57BL/6J)
Overall Breeding Performance
Good
Viability and Fertility: Female Male Comments
Homozygotes are viable: Undetermined Undetermined
Homozygotes are fertile: Undetermined Undetermined
Heterozygotes are fertile: Yes Yes
Age Reproductive Decline: Undetermined Undetermined
Average litter size
4 to 6
Recommended wean age
3 Weeks
Average Pups Weaned
4 to 6

Order Request Information

Limited quantities of breeder mice (recovered litter) are available from a cryoarchive; recovered litter usually available to ship in 3 to 4 months.

Cryopreserved material may be available upon request, please inquire to mmrrc@missouri.edu for more information.

Distribution of this strain requires submission of the MMRRC Conditions of Use (COU). A link to the COU web form will be provided via email after an order has been placed; the form should be completed then or the email forwarded to your institutional official for completion.

The donor or their institution limits the distribution to non-profit institutions only.

Additional charges may apply for any special requests. Shipping costs are in addition to the basic distribution/resuscitation fees. Information on shipping costs and any additional charges will be provided by the supplying MMRRC facility.

Click button to Request this one strain. (Use the MMRRC Catalog Search to request more than one strain.)
MMRRC Item # Description Distribution Fee / Unit (US $)
*Shipping & Handling not included*
Units Notes
068058-MU-SPERM Cryo-preserved spermatozoa $437.00 / Non-Profit Aliquot Approximate quantity3
068058-MU-RESUS Litter recovered from cryo-archive $2,624.00 / Non-Profit Litter Recovered litter4; additional fees for any special requests.
Cryopreserved material may be available upon request, please inquire to mmrrc@missouri.edu for more information.

1 The distribution fee covers the expense of rederiving mice from a live mouse; you will receive the resulting litter. The litter will contain at minimum one mutant carrier; the actual number of animals and the gender and genotype ratios will vary. (Typically, multiple breeder pairs can be established from the recovered litter.) Prior to shipment, the MMRRC will provide information about the animals recovered. If you anticipate or find that you need to request specific genotypes, genders or quantities of mice in excess of what is likely from a resuscitated litter, you may discuss available options and pricing with the supplying MMRRC facility.

2 An aliquot contains a sufficient number of embryos (in one or more vials or straws and based on the transfer success rate of the MMRRC facility) to transfer into one to three recipients. The MMRRC makes no guarantee concerning embryo transfer success experienced in the recipient investigator's laboratory. Neither gender nor genotype ratios are guaranteed.

3 An aliquot is one straw or vial with sufficient sperm to recover at least one litter of mice, as per provided protocols, when performed at the MMRRC facility. The MMRRC makes no guarantee concerning the success of these procedures when performed outside the MMRRC facilities.

4 The distribution fee covers the expense of resuscitating mice from the cryo-archive; you will receive the resulting litter. The litter will contain at minimum one mutant carrier; the actual number of animals and the gender and genotype ratios will vary. (Typically, multiple breeder pairs can be established from the recovered litter.) Prior to shipment, the MMRRC will provide information about the animals recovered. If you anticipate or find that you need to request specific genotypes, genders or quantities of mice in excess of what is likely from a resuscitated litter, you may discuss available options and pricing with the supplying MMRRC facility.

To request material from the MMRRC: Please fill out our on-line request form (accessible from the catalog search results page, or click the Request this Strain button in the fees section). If you have questions or need assistance completing this form, you may call Customer Service at (800) 910-2291 (in USA or Canada) or (530) 757-5710 (international calls). Before you call, please have with you: the MMRRC item number, quantity needed, Bill-to and Ship-to contact information.