Topic

How does Cyagen house its mice?

Cyagen’s top priority is to provide high quality mouse and rat models. Our facilities are accredited by the AAALAC (Association for Assessment and Accreditation of Laboratory Animal Care International), and are staffed with highly trained professionals. We pride ourselves on our ability to supply specific-pathogen free mice to animal facilities around the world. Features of the Cyagen Transgenic Animal Center include:

  • Specific-pathogen free (SPF) animal health status
  • Individually ventilated cage (IVC) and exhaust ventilated cage (EVC) systems that can house up to 60,000 mice
  • Class 10,000 ultra-clean environment
  • Quarterly sentinel rodent health monitoring program
  • Quarantine for all incoming rodents prior to colony integration

To see Cyagen’s list of excluded pathogens, please click here .

What animal strains does Cyagen use?

We typically produce custom animal models using the following strains, but we may be able to accommodate requests for other strains.

  • Transgenic mice: FVB or C57BL/6
  • Transgenic rats: Sprague-Dawley or Long Evans
  • Gene targeting (KO/KI): C57BL/6
  • Genome Editing: C57BL/6 or FVB (mice), Sprague-Dawley or Long Evans (rats)

Where does Cyagen source its animals?

We source our animals mainly from Charles River Laboratories. Currently, only mice from Charles River, Jackson Labs and Taconic Farms are allowed into our facility.

What is the difference between transgenics, knockouts, and knockins?

The key difference between transgenic and gene targeted (knockout or knockin) animals is that in knockout and knockin models the genomic changes are targeted, meaning a desired DNA sequence is altered at a specific locus in the target genome via homologous recombination, with an engineered segment of DNA replacing a homologous endogenous segment. By contrast, in transgenic animals an engineered segment of DNA is randomly integrated, such that the inserted DNA could end up almost anywhere in the host genome, and does not normally disrupt the homologous endogenous sequence.

Knockouts (KO) and knockins (KI) are both types of gene targeting, and differ from one another in the functional outcome for the targeted gene. Knockouts are changes that result in the inactivation of the targeted gene, such that the locus makes no gene product, or makes only a nonfunctional gene product. Knockins are targeted insertions into the genome that result in an altered gene product, such as a point mutation or the addition of a fluorescent tag. Knockout and Knockins are usually made using homologous recombination in ES cells, but nuclease-mediated genome editing using TALEN and CRISPR/Cas9 can also be used to generate knockouts or small sequence changes.

What is nuclease-mediated genome editing?

Artificially constructed nucleases (e.g., TALEN and CRISPR/Cas9) can be engineered to recognize and cleave arbitrary sequences. When such nucleases (or their DNA or mRNA precursors) designed to target a specific site in the mouse or rat genome are microinjected into fertilized eggs, cleavage at the target site followed by imperfect repair can result in small deletions (and insertions, more rarely) of one or more base pairs. If the cut site is in the coding region of a gene, this will result in frameshift mutations downstream of the site, generating a knockout. If a repair template with homology to the target site is present during the repair process, specific point mutations in the repair template can be introduced at the nuclease cleavage site, generating a knockin. Genome editing using TALEN or CRISPR/Cas9 can generate a knockout or knockin in as little as ~2 months, far faster than the typical 8-12 months required for conventional knockouts, because there is no need for ES cell work or chimera production.

What is included in your animal model services?

We offer all the services necessary to create your desired mouse or rat models. This includes free strategy design prior to your purchase, construction of transgenic or targeting vectors (including BAC recombineering), creating and characterizing targeted ES cells, microinjection of mouse or rat embryos, and breeding and genotyping of the resulting animals. For enhancer testing in mouse embryos using the LacZ reporter, we also offer LacZ staining of embryos.

How many embryos are injected for transgenic production, and what is the success rate?

Transgenic embryo production is carried out in sessions, with around 200 embryos injected per session. For plasmids, 3-4% (5-8) genotype-positive animals are typically obtained per session. For BACs, 1-2% (2-3) genotype-positive animals are typically obtained per session. These success rates are on par with, or better than, other transgenic service providers. Because the success rate for BACs is about half of that for plasmids, we recommend that our customers purchase two sessions for each BAC construct if they wish to obtain 4 or more genotype-positive animals per construct. Discounts are available for the multiple session of a construct.

Is it possible to customize procedures for fixing and LacZ-staining of transgenic embryos?

For enhancer testing in mouse embryos using the LacZ reporter, we offer a standard LacZ staining service of the embryos. However, we can also follow custom protocols provided by customers when fixing/staining embryos. For example, most customers have us perform LacZ staining, but some customers want us to only fix the embryos so they can do the staining themselves. It is also common for customers to request the dissection of specific organs for LacZ staining.

Would lacZ staining interfere with antibody-based staining of protein of interest?

We are not entirely sure, but we suspect that lacZ staining might interfere with antibody staining. If the customer really wants to see lacZ staining and antibody staining in the same embryo, we would suggest dividing the embryo into two left and right halves, and use one half for lacZ staining and the other half for antibody staining. For antibody staining, it may have to be done on sections, as we have never seen whole-mount antibody staining.

How many embryos would you be able to send us and in which period of time?

4 transgene positive embryos (based on PCR genotyping) with guaranteed service for plasmid injection, but typically more transgene positive embryos are actually obtained. The customer can request for guarantee of more than 4 embryos, but the price would be adjusted accordingly.

How are the embryos collected and delivered?

Embryos are isolated from surrogate mothers at the appropriate embryonic age through standard dissection from uterus, with care taken to minimize any damage to the embryo. Embryos are then fixed in 4% PFA (for 1-3 hours based on embryonic age). If lacZ staining is required, then it is performed by standard whole-mount staining protocol.Embryos or organs, whether fixed only or fixed followed by lacZ stained, are shipped in PBS solution at room temperature in tubes of the appropriate size.

Where can Cyagen ship to?

Cyagen can ship to many countries worldwide. We have successfully delivered custom animal models and other products to over a dozen countries, including Australia and many European and Asian countries. Some countries have specific regulations regarding the shipment of biological materials, but we will work with you to deal with these issues.

How does Cyagen ship embryos to customers?

We ship embryos under ambient temperature by express carrier. It typically takes 3 work days to reach anywhere in the world. This has worked well for all our customers, whether they have us do LacZ staining or perform their own staining after receiving fixed embryos from us.

How should customers ship DNA to Cyagen?

Customers can ship plasmid or BAC DNA to us by an express carrier under ambient temperature. In our experience, DNA quality is not compromised by this shipping method if the DNA is prepared according to our recommendation. We will test DNA quality before use and inform customers of any signs of quality problems.

How long time would it take from sending the order to delivering of the embryos?

Around 1.5-3 months, depending on whether vector construction is needed. If there is no vector construction, it should be around 1.5 months in most cases.

Who do I contact if I have a question about my order?

For answers to questions about an animal order that you have placed, please contact your assigned Cyagen project manager via email. You should have received an email from your project manager when your animal project was initiated.

For other questions, including technical questions, please submit a message online, send us an email at service@cyagen.com, or call us at 800-921-8930.

Why haven’t I received update emails or replies from Cyagen?

Email spam filters occasionally place Cyagen correspondence in spam folders. Please check your spam folder and add Cyagen email addresses to your address book to ensure that our emails are properly delivered.

What payment methods can customers use?

US clients must remit payment by check, wire transfer or credit/debit card. Non-US clients must remit payment by wire transfer or credit/debit card. Wire transfers may involve fees charged directly to the transferred amount by originating and/or intermediate banks. To avoid underpayment, please discuss with your bank in advance to ensure transfer of sufficient funds such that the net value of your wire transfer, after all fees charged by originating and intermediate banks, is equal to your outstanding invoice balance.

Check (for US buyers):

Payable to: Cyagen Biosciences Inc.
Mail check to: Cyagen Biosciences Inc. 2255 Martin Avenue, Suite E Santa Clara, CA 95050, USA

Wire transfer:

Beneficiary: Cyagen Biosciences Inc.
Beneficiary Bank Name: JPMorgan Chase Bank, N.A
Beneficiary Bank Routing Number: 322271627
Beneficiary Bank Account Number: 573797979
Beneficiary Bank Swiss Code: CHASUS33
Beneficiary Bank Address: 2200 El Camino Real Santa Clara, CA 95050 USA

Credit/debit card:

Please contact us at 800-921-8930 or accounting@cyagen.com to process credit card payment.

How does Cyagen protect customers' Intellectual Property (IP)?

All materials from our customers, such as DNA constructs, PCR primers, and cells, are coded internally once they reach us, and kept strictly within the company’s premises. All information and experimental results of customers’ projects are maintained in secure places and kept apart from all other projects. Only authorized personnel have access to these materials and information.