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Each full transgenic project is split into the following phases:
1. Transgenic strategy design
2. Transgenic vector construction
3. Pronuclear injection
4. Transgenic mouse screening
Preparation of DNA for injection and development of a genotyping strategy
The success of transgenic production depends critically on the quality of DNA used in pronuclear injection. Accordingly, we employ a special protocol to prepare top-quality DNA for our injections. Plasmid DNA is typically linearized for injection. BAC DNA is typically injected in unlinearized form, as this has been shown to produce similar biological outcomes as linearized BAC. A PCR-based genotyping strategy will be developed and tested. If you provide your own transgenic vector rather than having us construct it, you can send us either E. coli or unlinearized DNA in solution (≥50 µg total), and we will perform the necessary preparation to purify/linearize the DNA. We strongly discourage customers from sending us linearized, injection-ready DNA, because the quality of such DNA is often insufficient for pronuclear injection. If you provide your own vector without sequence information, you will also need to provide a PCR-based genotyping strategy along with your vector.
Generation of transgenic founder mice
- Plasmid or BAC injection to obtain founders, basic service: A minimum of 400 eggs (C57BL/6) will be injected to produce live animals. This will result in an average of 2-3 transgenic founders in the case of plasmid injection or 1-2 founders in the case of BAC injection, but the actual numbers could fluctuate significantly and it is possible that no founders are obtained. If you wish to ensure the generation of founders, we strongly recommend that you use our guaranteed service option (see below).
- Plasmid or BAC injection to obtain founders, guaranteed service: We guarantee a minimum of 3 transgenic founders for plasmid injection or 2 founders for BAC injection (the actual number of founders obtained is typically higher).
- Genotyping pups to identify founders: Pups will be genotyped by PCR to identify founder mice.
We typically produce transgenic mice in C57BL/6 strain backgrounds, but we can use other strains per your request.
|Service||Donor egg strain||Price||Turnaround time|
|Transgenic strategy design||Free||1-4 days|
|Transgenic vector construction or BAC modification*||Please inquire||2-5 weeks|
|Prepare DNA for injection and develop genotyping strategy||$500||1-2 weeks|
|Plasmid injection to obtain founders, basic service||C57BL/6||$3,450||6-10 weeks|
|Plasmid injection to obtain founders, guaranteed service||C57BL/6||$3,950||6-16 weeks|
|BAC injection to obtain founders, basic service*||C57BL/6||$4,450||8-14 weeks|
|BAC injection to obtain founders, guaranteed service*||C57BL/6||$6,950||8-18 weeks|
|Genotyping pups to identify founders||$250||1 week|
Cyagen offers the best guarantee in the industry – we will fully refund the client’s service fee if animals with the specified genotype are not generated (except for genetic modifications severely affecting viability, morbidity, or fertility). Given the complexity of biological systems, a particular genetic modification may not result in the desired phenotype. As such, Cyagen's guarantee covers the creation of animals with the specified genotype, not a particular phenotypic outcome in terms of transcription, protein/RNA function, or organismal biology.
If you find another commercial service provider that offers better pricing than ours, we will match the price plus an additional 5% off.
Standard payment terms include a 50% upfront payment before the project, and the remaining 50% plus shipping charge paid after completion of the project. If you need us to design your transgenic strategy, we will provide this service for free irrespective of whether you end up choosing us for your project.
We offer up to a 10% bulk discount for large orders. Large orders are defined as 5 or more projects from the same institution. If you bundle your orders with those of your colleagues, you can all qualify for the bulk discount.
Products are shipped from our Santa Clara, California facility. For mouse shipments, the shipping charge includes courier cost plus a $100/crate handling fee. DNA constructs or fixed/stained embryos are shipped at room temperature, and the charge includes courier cost plus a $10 handling fee. We typically use World Courier to ship live mice and FedEx for other shipments.
All animal work is conducted in our specific pathogen free (SPF) facilities that have been AAALAC accredited and OLAW assured. For details information, please visit our support section for Description of our Facility, Animal Health and Animal Welfare Program.
Please click here to view a map of customers who have used Cyagen before worldwide.
Please click here for a list of publications that have cited Cyagen.
◆ Case studies on our Regular Transgenic Mice
Cardiac Fibroblast-Specific Activating Transcription Factor 3 Protects Against Heart Failure by Suppressing MAP2K3-p38 Signaling.
Circulation 135: 2041-2057 (2017)
Li Y, Li Z, Zhang C, Li P, Wu Y, Wang C, Bond Lau W, Ma XL, Du J
BACKGROUND: Hypertensive ventricular remodeling is a common cause of heart failure. However, the molecular mechanisms regulating ventricular remodeling remain poorly understood. METHODS: We used a discovery-driven/nonbiased approach to identify increased activating transcription factor 3 (ATF3) expression in hypertensive heart. We used loss/gain of function approaches to understand the role of ATF3 in heart failure. We also examined the mechanisms through transcriptome, chromatin immunoprecipitation sequencing analysis, and in vivo and in vitro experiments. RESULTS: ATF3 expression increased in murine hypertensive heart and human hypertrophic heart. Cardiac fibroblast cells are the primary cell type expressing high ATF3 levels in response to hypertensive stimuli. ATF3 knockout (ATF3KO) markedly exaggerated hypertensive ventricular remodeling, a state rescued by lentivirus-mediated/miRNA-aided cardiac fibroblast-selective ATF3 overexpression. Conversely, conditional cardiac fibroblast cell-specific ATF3 transgenic overexpression significantly ameliorated ventricular remodeling and heart failure. We identified Map2K3 as a novel ATF3 target. ATF3 binds with the Map2K3 promoter, recruiting HDAC1, resulting in Map2K3 gene-associated histone deacetylation, thereby inhibiting Map2K3 expression. Genetic Map2K3 knockdown rescued the profibrotic/hypertrophic phenotype in ATF3KO cells. Last, we demonstrated that p38 is the downstream molecule of Map2K3 mediating the profibrotic/hypertrophic effects in ATF3KO animals. Inhibition of p38 signaling reduced transforming growth factor-β signaling-related profibrotic and hypertrophic gene expression, and blocked exaggerated cardiac remodeling in ATF3KO cells. CONCLUSIONS: Our study provides the first evidence that ATF3 upregulation in cardiac fibroblasts in response to hypertensive stimuli protects the heart by suppressing Map2K3 expression and subsequent p38-transforming growth factor-β signaling. These results suggest that positive modulation of cardiac fibroblast ATF3 may represent a novel therapeutic approach against hypertensive cardiac remodeling.
Chromodomain Protein CDYL Acts as a Crotonyl-CoA Hydratase to Regulate Histone Crotonylation and Spermatogenesis.
Experimental cell research 67: 853-866.e5 (2017)
ng Y, Liu S, Yu H, Liu Y, Liu X
Lysine crotonylation (Kcr) is a newly identified histone modification that is associated with active transcription in mammalian cells. Here we report that the chromodomain Y-like transcription corepressor CDYL negatively regulates histone Kcr by acting as a crotonyl-CoA hydratase to convert crotonyl-CoA to β-hydroxybutyryl-CoA. We showed that the negative regulation of histone Kcr by CDYL is intrinsically linked to its transcription repression activity and functionally implemented in the reactivation of sex chromosome-linked genes in round spermatids and genome-wide histone replacement in elongating spermatids. Significantly, Cdyl transgenic mice manifest dysregulation of histone Kcr and reduction of male fertility with a decreased epididymal sperm count and sperm cell motility. Our study uncovers a biochemical pathway in the regulation of histone Kcr and implicates CDYL-regulated histone Kcr in spermatogenesis, adding to the understanding of the physiology of male reproduction and the mechanism of the spermatogenic failure in AZFc (Azoospermia Factor c)-deleted infertile men.