The purpose of the Genetically Engineered Mouse Facility (GEMF) is to provide technologically advanced and efficient mouse mutation resources to faculty members at the institution. To modify the genome, we perform direct DNA injection or electroporation for standard transgenesis and CRISPR-mediated gene targeting. We also perform ES cell mutagenesis followed by blastocyst injection to generate modifications in the classical manner. Archiving and assisted reproduction, including embryo and sperm cryopreservation, in vitro fertilization and rederivation of mouse lines are all technologies supported by the GEMF.
The facility is fully equipped with the latest instrumentation and staffed with highly skilled personnel trained specifically in the production of mutant mice.
Revised Expectations for CRISPR-Mediated Generation of Conditional Alleles
The GEMF and several other MD Anderson investigators participated in a worldwide consortium to investigate the ease with which conditional alleles could be created using 2 sgRNAs and 2 donor oligos, as published in 2012 by Yang, et al [Cell, 2013. 153(4): 910-18]. An analysis of 17,887 injected or electroporated zygotes resulted in 1,718 live born mice. Of these only 15 had the correct, two loxP insertion sites for an overall rate of <1%. The manuscript describing this study is now available Gurumurthy C et al. Biol. 2019 Aug 26;20(1):171. doi: 10.1186/s13059-019-1776-2.PMID: 31446895
New Injection Criteria for CRISPR/Cas9 Injections
Based on our current experience with CRISPR/Cas9 projects, and due to the availability of high quality components, we are now adjusting our injection structure for CRISPR gene targeting projects. For CRISPR knock-out projects, we will perform two days of injection. For CRISPR knock-in projects, or projects based on hdr, we will perform 3 days of injections. We cannot guarantee success, but an investigator may request additional injection days at a standard cost per day. Please see our Services and Fees section for more details.
The Dead Live Again
Did the last male of your critical line just die? If you put him in the refrigerator immediately (NOT the freezer), we can isolate sperm to recover the line for up to four days after death. In a previous procedure using a 6 to 8 month old male that had unfortunately expired, we ended up with a 79% fertilization rate. So don't give up hope. Call us at 713-745-1419 or 713-745-2654.
Sperm Cryopreservation and Assisted Reproduction
The GEMF uses the sperm cryopreservation and IVF protocols developed by Naomi Nakagata at the CARD Institute in Japan. To archive your animals, you may request sperm cryopreservation, embryo cryopreservation, or an IVF-embryo cryopreservation. Please see our Services and Fees section for more details.