The MADM-7TG (Hipp7TG) allele has the CMV enhancer/chicken beta-actin core promoter, the N-terminal portion of tdTomato, a beta-globin intronic sequence (containing frt and lox sites), and the C-terminal portion of mut4-EGFP all inserted into the Hipp7 locus on chromosome 7 (~2.13 cM; ~0.7 kbp downstream of exon 5 of the Rps9 gene). These MADM-7TG mutants are designed for MADM (mosaic analysis with double markers), and must be crossed to MADM-7GT mice harboring a reciprocal mutation at the same locus (Stock No. 021457). This MADM system allows Cre recombinase-induced fluorescent labeling of daughter cells to ascertain lineal relationships and pleiotropic gene function in multicellular organisms. These mice may also be useful in studies of cell differentiation, mitosis, and imprinting.
Simon Hippenmeyer, IST Austria (Institute of Science and Technology Austria)
Dr. Liqun Luo, Stanford University, HHMI
Mice homozygous for the MADM-7TG (Hipp7TG) allele are viable and fertile with no reported abnormalities. The MADM-7TG allele has the CMV enhancer/chicken beta-actin core promoter, the N-terminal portion of tdTomato, a beta-globin intronic sequence (containing a frt-flanked region of two lox site variants [lox2272 and lox5171] and then a loxP-flanked neomycin resistance gene), and the C-terminal portion of mut4-EGFP all inserted into the Hipp7 locus on chromosome 7 (~2.13 cM; ~0.7 kbp downstream of exon 5 of the Rps9 gene). These MADM-7TG mutants are designed for MADM (mosaic analysis with double markers), and must be crossed to MADM-7GT mice harboring a reciprocal mutation at the same locus (Stock No. 021457). The resulting TG/GT offspring have one copy of each reciprocal mutation on homologous chromosomes ("trans-heterozygous"), and must also be bred to harbor a Cre- or FLP-recombinase to induce fluorescent protein expression. Prior to Cre- or FLP-recombination, trans-heterozygous mutant mice do not have colored cells: the chimeric genes do not produce functional proteins because their coding sequences are interrupted by the beta-globin intron in different reading frames. After DNA replication (G2 phase) in double mutant mice, introduction of Cre- or FLP-recombinase that facilitates inter-chromosomal recombination aligns the respective N- and C-terminal coding sequences for each of the reporter genes on the same chromosome. The subsequent chromatid segregation (X or Z) determines daughter cell phenotype: recombinant sister chromatids into the same daughter cell (a G2-Z event) leads to double reporter expression or no reporter expression, while independent segregation into separate daughter cells (a G2-X event) leads to expression of either EGFP or tdTomato-MYC. If an additional targeted mutation of interest is introduced distal to the Hipp7 locus on chromosome 7, only homozygous cells will be singly labeled following G2 cre or FLP introduction. The homozygous mutant and wildtype cells can then be distinguished by which single reporter they express. Most heterozygous cells will be unlabeled, but some heterozygous cells will be yellow (both markers expressed). Reporter protein tissue specificity, expression levels, and frequency of recombination are thus determined by the promoter controlling Cre- or FLP-recombinase expression. Using this MADM system, a researcher can generate genetic mosaics in which an individual organism contains somatic cells of different genotypes. This allows the researcher to ascertain lineal relationships and pleiotropic gene function in multicellular organisms. These mice may also be useful in studies of cell differentiation and mitosis.
Other important features of the MADM-7 system are listed below. Because of its placement ~3.6 Mbp from the centromere, MADM-7 allows almost all of the genes on chromosome 7 to be subjected to MADM-based mosaic analyses. Cre- or FLP-recombinase introduction in cell phase G0 or G1 results in double reporter expression. The donating investigator also reports the MADM-7 design has several advantages compared to the original MADM(-6) mice. Specifically, MADM-7 allows direct fluorescent visualization of both EGFP and tdTomato in live animals/cells: permitting genotypes of distinctly labeled cells in mosaic animals to be unequivocally determined prior to fixation and/or immunostaining. Also, MADM-7 contains both loxP and frt sites; allowing the induction of MADM-labeling by either Cre recombinase or FLP recombinase.
The same donating investigator has several mice with MADM applications on different chromosomes:
On chromosome 6, the Gt(ROSA)26Sor knockin mutations include MADM-6GR (Stock Nos. 006041 / 006075), MADM-6RG (Stock Nos. 006067 / 006080),MADM-6GG (Stock Nos. 006053 / 006071), R26GT (Stock No. 017912), R26TG (Stock No. 017921), R26TT (Stock No. 017922), and R26G-tTA2 (Stock No. 017909).
On chromosome 7, the centromeric insertions are MADM-7GT (Hipp7GT; Stock No. 021457) andMADM-7TG (Hipp7TG; Stock No. 021458).
On chromosome 10, the centromeric insertions are MADM-10GT (Miya10GT; Stock No. 017923) andMADM-10TG (Miya10TG; Stock No. 017932).
On chromosome 11, the centromeric insertions are MADM-11GT (Hipp11GT; Stock No. 013749) andMADM-11TG (Hipp11TG; Stock No. 013751).
On chromosome 12, the centromeric insertions are MADM-12GT (John12GT; Stock No. 021460) andMADM-12TG (John12TG; Stock No. 021461).
A targeting vector was created with a CMV enhancer/chicken beta-actin core promoter (pCA) upstream of the "TG MADM" cassette. The "TG MADM" cassette used here was designed with the N-terminal portion of a red fluorescent protein (tdTomato; aa 1-3), a beta-globin intronic sequence (containing a frt-flanked region of two lox site variants [lox2272 and lox5171] and then a loxP-flanked neomycin resistance gene), the C-terminal portion of a mutant enhanced green fluorescent protein (mut4-EGFP; aa 274-724), and an SV40 T-antigen poly(A) signal. This entire "TG MADM" construct was inserted into the Hipp7 locus on chromosome 7 (~2.13 cM; ~0.7 kbp downstream of exon 5 of the Rps9 gene) via electroporation of (129X1/SvJ x 129S1/Sv)F1-derived R1 embryonic stem (ES) cells. Correctly targeted ES cells were microinjected into recipient blastocysts. Chimeric males were bred to CD1 females to establish the MADM-7TG colony. The donating investigator also reports that MADM-7TG mice were previously bred to Emx1-Cre mice (non-germline Cre strain on an undisclosed genetic background), and the Cre recombinase was later removed. MADM-7TG mice on a mixed CD1;129;C57BL/6 genetic background (white coat color) were sent to The Jackson Laboratory Repository in 2013. Upon arrival, sperm was cryopreserved. To generate the living colony, an aliquot of the frozen sperm was used to fertilize oocytes from C57BL/6J female mice (Stock No. 000664). The MADM-7TG mice were then bred together, to wildtype mice from the colony, or to C57BL/6J mice.
|Allele Name||targeted mutation 1, Liqun Luo|
|Allele Type||Targeted (Conditional ready (e.g. floxed), No functional change)|
|Allele Synonym(s)||Iis5tm1(ACTB-tdTomato,-EGFP)Luo; MADM-7TG|
|Gene Symbol and Name||Igs5, intergenic site 5|
|Gene Synonym(s)||Hipp7; Iis5; MADM-7|
|Strain of Origin||(129X1/SvJ x 129S1/Sv)F1-Kitl<+>|
The average number of mice provided from recovery of our cryopreserved strains is 10. The total number of animals provided,
their gender and genotype will vary. We will fulfill your order by providing at least two pair of mice, at least one animal of
each pair carrying the mutation of interest. Please inquire if larger numbers of animals with specific genotype and genders
are needed. Animals typically ship between 10 and 14 weeks from the date of your order. If a second cryorecovery is needed in
order to provide the minimum number of animals, animals will ship within 25 weeks.
The genotypes of animals provided may not reflect the mating scheme utilized by The Jackson Laboratory prior to cryopreservation, or that discussed in the strain description. Please inquire about possible genotypes which will be recovered for this specific strain. The Jackson Laboratory 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.
Terms are granted by individual review and stated on the customer invoice(s) and account statement. These transactions are payable in U.S. currency within the granted terms. Payment for services, products, shipping containers, and shipping costs that are rendered are expected within the payment terms indicated on the invoice or stated by contract. Invoices and account balances in arrears of stated terms may result in The Jackson Laboratory pursuing collection activities including but not limited to outside agencies and court filings.
The Jackson Laboratory has rigorous genetic quality control and mutant gene genotyping programs to ensure the genetic background of JAX® Mice strains as well as the genotypes of strains with identified molecular mutations. JAX® Mice strains are only made available to researchers after meeting our standards. However, the phenotype of each strain may not be fully characterized and/or captured in the strain data sheets. Therefore, we cannot guarantee a strain's phenotype will meet all expectations. To ensure that JAX® Mice will meet the needs of individual research projects or when requesting a strain that is new to your research, we suggest ordering and performing tests on a small number of mice to determine suitability for your particular project.
"MICE" means mouse strains, their progeny derived by inbreeding or crossbreeding, unmodified derivatives from mouse strains or their progeny supplied by The Jackson Laboratory ("JACKSON"). "PRODUCT(S)" means biological materials supplied by JACKSON, and their derivatives. "SERVICES" means projects conducted by JACKSON for other parties that may include but are not limited to the use of MICE or PRODUCTS. "RECIPIENT" means each recipient of MICE, PRODUCTS, or SERVICES provided by JACKSON including each institution, its employees and other researchers under its control. MICE or PRODUCTS shall not be: (i) used for any purpose other than internal research, (ii) sold or otherwise provided to any third party for any use, or (iii) provided to any agent or other third party to provide breeding or other services. Acceptance of MICE, PRODUCTS or SERVICES from JACKSON shall be deemed as agreement by RECIPIENT to these conditions, and departure from these conditions requires JACKSON’s prior written authorization.
MICE, PRODUCTS AND SERVICES ARE PROVIDED "AS IS". JACKSON EXTENDS NO WARRANTIES OF ANY KIND, EITHER EXPRESS, IMPLIED, OR STATUTORY, WITH RESPECT TO MICE, PRODUCTS OR SERVICES, INCLUDING ANY IMPLIED WARRANTY OF MERCHANTABILITY OR FITNESS FOR A PARTICULAR PURPOSE, OR ANY WARRANTY OF NON-INFRINGEMENT OF ANY PATENT, TRADEMARK, OR OTHER INTELLECTUAL PROPERTY RIGHTS.
In case of dissatisfaction for a valid reason and claimed in writing by a purchaser within ninety (90) days of receipt of, PRODUCTS or SERVICES, JACKSON will, at its option, provide credit or replacement for the PRODUCT received or the SERVICES provided; JACKSON makes no other representations and this shall be the exclusive remedy of the purchaser. Please note specific policy for live mice.
Consistent with the requirement for a written understanding regarding animal care and use, the JACKSON Animal Care and Use Committee will review the animal care and use protocol(s) associated with any SERVICES to be performed at JACKSON, and JACKSON shall have ultimate responsibility and authority for the care of animals while on site or in JACKSON custody.
In no event shall JACKSON, its trustees, directors, officers, employees, and affiliates be liable for any causes of action or damages, including any direct, indirect, special, or consequential damages, arising out of the provision of MICE, PRODUCTS, or SERVICES, including economic damage or injury to property and lost profits, and including any damage arising from acts or negligence on the part of JACKSON, its agents or employees. Unless prohibited by law, in purchasing or receiving MICE, PRODUCTS, or SERVICES from JACKSON, purchaser or recipient, or any party claiming by or through them, expressly releases and discharges JACKSON from all such causes of action or damages, and further agrees to defend and indemnify JACKSON from any costs or damages arising out of any third party claims.
MICE, PRODUCTS or SERVICES are to be used in a safe manner and in accordance with all applicable governmental rules and regulations.
The foregoing represents the General Terms and Conditions applicable to JACKSON’s MICE, PRODUCTS or SERVICES. In addition, special terms and conditions of sale of certain MICE, PRODUCTS, or SERVICES may be set forth separately in JACKSON web pages, catalogs, price lists, contracts, and/or other documents, and these special terms and conditions shall also govern the sale of these MICE, PRODUCTS and SERVICES by JACKSON, and by its licensees and distributors.
Acceptance of delivery of MICE, PRODUCTS or SERVICES shall be deemed agreement to these terms and conditions. No purchase order or other document transmitted by purchaser or recipient that may modify the terms and conditions hereof, shall be in any way binding on JACKSON, and instead the terms and conditions set forth herein, including any special terms and conditions set forth separately, shall govern the sale of MICE, PRODUCTS or SERVICES by JACKSON.