To generate MMTV-Cre; mPot1aF/F; p53F/F mice, MMTV-Cre mice (The Jackson Laboratory, Tg (MMTV-cre) 4Mam/J; stock number: 003553) were first crossed with mPot1aF/F mice (Wu et al., 2006 (link)) to generate MMTV-Cre; mPot1aF/F mice. MMTV-Cre; mPot1aF/F mice were then crossed with p53F/F (The Jackson Laboratory, B6.129P2-Trp53tm1Brn/J; Stock No:008462) mice to generate MMTV-Cre; mPot1aF/F; p53F/+ and MMTV-Cre; mPot1aF/F; p53F/F mice. mPot1aF/F and mPot1aF/F; p53F/F mice were generated as controls. All mice were maintained under basic care conditions according to the IACUC-approved protocols of Yale University. Sick mice (age 7–12 months old; Table S1 ) were sacrificed and tumors were harvested from breast glands or other organs. Chopped tumors were digested with 0.25% trypsin at 37° C for 15min followed by collagenase D treatment at 37° C for 30 min. The digested suspension was filtered through 40μm cell strainer and isolated tumor cells were pelleted by centrifugation and expanded by passaging in the DMEM with 10% FBS culture media.
Mmtv cre mice
Manufactured by Jackson ImmunoResearch
MMTV-Cre mice are a strain of transgenic mice that express the Cre recombinase enzyme under the control of the mouse mammary tumor virus (MMTV) promoter. The Cre recombinase enzyme can be used to conditionally delete or activate target genes in specific cell types, such as mammary epithelial cells, during the lifetime of the mouse.
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4 protocols using mmtv cre mice
1
Generating MMTV-Cre; mPot1aF/F; p53F/F Mice
2
Generating MMTV-Cre; mPot1aF/F; p53F/F Mice
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To generate MMTV-Cre; mPot1aF/F; p53F/F mice, MMTV-Cre mice (The Jackson Laboratory, Tg (MMTV-cre) 4Mam/J; stock number: 003553) were first crossed with mPot1aF/F mice (Wu et al., 2006 (link)) to generate MMTV-Cre; mPot1aF/F mice. MMTV-Cre; mPot1aF/F mice were then crossed with p53F/F (The Jackson Laboratory, B6.129P2-Trp53tm1Brn/J; Stock No:008462) mice to generate MMTV-Cre; mPot1aF/F; p53F/+ and MMTV-Cre; mPot1aF/F; p53F/F mice. mPot1aF/F and mPot1aF/F; p53F/F mice were generated as controls. All mice were maintained under basic care conditions according to the IACUC-approved protocols of Yale University. Sick mice (age 7–12 months old; Table S1 ) were sacrificed and tumors were harvested from breast glands or other organs. Chopped tumors were digested with 0.25% trypsin at 37° C for 15min followed by collagenase D treatment at 37° C for 30 min. The digested suspension was filtered through 40μm cell strainer and isolated tumor cells were pelleted by centrifugation and expanded by passaging in the DMEM with 10% FBS culture media.
3
Conditional Knockout Mice for mTOR Pathway
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All animals were housed under pathogen-free conditions, and experiments were performed in accordance with AAALAC guidelines and with Vanderbilt University Institutional Animal Care and Use Committee approval. RictorFL/FL mice (C57BL/6) were kindly provided by Dr. Mark Magnuson (Vanderbilt University) and have been previously described [14 (link)]. RaptorFL/FL mice ([29 (link)], C57BL/6) were purchased from the Jackson Laboratories (Bar Harbor, ME). MMTV-Cre mice ([13 (link)] FVB) were purchased from the Jackson Laboratories. All analyses were performed on age-matched siblings resulting from F1 (1:1, FVB:C57BL/6) intercrosses.
4
Generation and Characterization of PARP1/2 KO Mice
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PARP1 and PARP2 global KO mice, as well as PARP2 flox mice were provided by Dr. Francoise Dantzer5 (link),19 (link),27 (link). Generation of PARP1 flox mice has been published28 (link). To establish conditional PARP1 or PARP2 KO in the myeloid lineage, PARP1 or PARP2 flox mice were bred with LyzM-Cre mice29 (link). MMTV-PyMT transgenic C57BL/6 mice and MMTV-Cre mice were from the Jackson Laboratory. WT C57BL/6 J mice (UTSW Breeding Core), BALB/cJ mice (UTSW Breeding Core), and athymic nude mice (Charles River) were also used. Nude PARP1 and PARP2 global KO mice were established by breeding PARP1 and PARP2 global KO C57BL/6 mice with athymic nude mice. All experiments were conducted using littermates. All protocols for mouse experiments were approved by the Institutional Animal Care and Use Committee of University of Texas Southwestern Medical Center. All mice were housed in accordance with approved IACUC protocols. Animals were housed on a 12–12 light cycle (light on 6:00, off 18:00) and provided food and water ad libitum.
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